,1,2, 李婉杰1, 李九丽1, 张弛1, 胡梦辉1, 程琳1, 袁红雨1,2The Research Progress of Plant RNA Binding Proteins
ZHANG ZaiBao,1,2, LI WanJie1, LI JiuLi1, ZHANG Chi1, HU MengHui1, CHENG Lin1, YUAN HongYu1,2通讯作者:
责任编辑: 李莉
收稿日期:2018-06-4接受日期:2018-08-8网络出版日期:2018-11-01
| 基金资助: |
Received:2018-06-4Accepted:2018-08-8Online:2018-11-01
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张在宝, 李婉杰, 李九丽, 张弛, 胡梦辉, 程琳, 袁红雨. 植物RNA结合蛋白研究进展[J]. 中国农业科学, 2018, 51(21): 4007-4019 doi:10.3864/j.issn.0578-1752.2018.21.001
ZHANG ZaiBao, LI WanJie, LI JiuLi, ZHANG Chi, HU MengHui, CHENG Lin, YUAN HongYu.
转录及转录后水平的基因表达调控对真核生物生长发育和响应外部环境刺激至关重要。真核生物基因表达的转录后调控包括RNA加工、前体RNA剪接、RNA的细胞核输出、RNA的稳定、翻译和凋亡,这些过程被称为RNA代谢。在RNA代谢途径中,大量的蛋白质因子——RNA结合蛋白（RNA-binding proteins,RBPs）与RNA结合形成核糖核蛋白（ribonucleoprotein,RNP）复合物来调节RNA代谢过程。每一种RNA分子都有许多RNA结合蛋白与之结合来稳定、保护、组装或转移它[1,2]。RNA结合蛋白具有多个保守的基序和结构域,包括RNA识别基序（RNA recognition motif,RRM）、锌指基序、K同源性结构域（K-homology domain,KH）、冷休克结构域、富含甘氨酸的区域、富含精氨酸的区域、RD重复序列和SR重复序列,这些结构域相互结合构成了各种特异性RBPs[3,4]。
RNA结合蛋白广泛存在于动物、植物以及微生物中,约占真核生物基因编码蛋白的2%—8%[5]。根据蛋白结构,RNA结合蛋白可以分为7个蛋白家族：富含丝氨酸-精氨酸的RNA结合蛋白（SR蛋白）、富含甘氨酸的RNA结合蛋白（GR-RBPs）、三角状五肽重复区蛋白（PPR蛋白）、DEAD-box RNA解旋酶（DEAD-box RHs）、叶绿体RNA剪接与核糖体成熟结构域蛋白（chloroplast RNA splicing and ribosome maturation domain protein,CRM蛋白）、S1结构域蛋白（S1 domain containing-protein,SDPs）和细菌冷休克蛋白（bacterial cold shock protein,CSPs）。在哺乳动物中,RNA结合蛋白与人类疾病如肿瘤、神经系统疾病等密切相关。例如RNPC1（RRM家族的RBP）通过调节癌细胞中mRNA的稳定性而实现靶标的差异表达,其异位表达会诱导细胞周期停滞,抑制乳腺肿瘤细胞增殖,并通过调节mutp53诱导的逆转上皮-间质转化（epithelial-mesenchymal transition,EMT）进一步抑制肿瘤细胞的迁移和侵袭[6]。TDP-43是核糖核蛋白（Heterogeneous nuclear ribonulcleoprotein,hnRNP）家族成员,其存在于海马神经元的树突中并能够结合数千个RNA分子,参与miRNA代谢,对神经元的生长不可或缺[7]。植物的RNA结合蛋白在植物生长发育以及应激反应过程中发挥重要作用,目前对RNA结合蛋白在植物中的功能和作用机制的研究还比较少。前人对植物RBPs的研究主要集中在核编码的靶向叶绿体或线粒体的RBPs、富含甘氨酸的RBPs（GR-RBPs）以及三角状五肽重复区蛋白（PPR蛋白）。大量的研究表明植物RBPs作为重要的调控因子在RNA代谢、植物生长发育以及胁迫响应中发挥重要作用,从而引起人们对植物RBPs的广泛关注。本研究阐述了一些与植物（主要是拟南芥）生长发育以及胁迫响应相关的RBPs的功能、作用机制和相互联系,重点总结了5类RBP家族在植物生长发育和胁迫反应中的重要作用（表1和表2）,包括SR蛋白、GR-RBPs、PPR蛋白、DEAD-box RHs以及RNA分子伴侣。
Table 1
表1
表1植物RNA结合蛋白的分类、结构特征及主要功能
Table 1
| 分类 Classification | 结构特征 Structural characteristics | 生理功能 Physiological function |
|---|---|---|
| SR蛋白 SR proteins | N端：含1个或2个RRM结构域; C端：富含SR的结构域 N-terminal: Containing 1 or 2 RRM domain; C-terminal: SR-rich domain | 参与mRNA剪接、输出或翻译,调节生长发育及细胞增殖;响应高温和低温等非生物胁迫 Participating in mRNA splicing, export or translation, regulating plant growth, development and cell proliferation; Responding to abiotic stresses such as heat and cold stresses |
| GR-RBPs (Ⅳ of GRPs） | N端：含RRM或CSD结构域; C端：存在富含甘氨酸区域 N-terminal: Containing RRM or CSD domain; C-terminal: Containing the regions of glycine rich | 作为RNA分子伴侣,响应干旱、高盐和低温等胁迫;调节植物生物钟,介导开花等过程 As RNA molecular chaperone, responding to drought, salt and cold stresses; Regulating plant circadian clock, mediating plants flowering, etc. |
| RZs (IVb of GRPs) | N端：含RRM结构域; C端：富含甘氨酸区域且其间散布CCHC型锌指基序 N-terminal: Containing RRM domain; C-terminal: The region rich in glycine and interspersed with CCHC-type zinc finger motifs | 响应低温和干旱等胁迫;与SR等蛋白互作参与mRNA剪接,促进种子萌发、幼苗生长和开花等植物发育 Responding to cold and drought stresses; interacting with SR and other proteins to participate in mRNA splicing, promote seed germination, seedling growth and flowering, etc. |
| CSDPs (IVc of GRPs) | N端：含一个CSD结构域; C端：含一个富含甘氨酸区域且其间散布CCHC型锌指基序 N-terminal: Contains a CSD domain; C-terminal: Containing a glycine-rich region interspersed with CCHC-type zinc finger motifs | 正/负调控植物的胁迫耐受性、生长发育等 Positively/negatively regulating the plant stress tolerance, growth and development, etc. |
| PPR蛋白 PPR proteins | 由大约35个氨基酸序列组成的串联重复基序 The tandem repeat motif consisting of approximately 35 amino acid sequences | 主要参与线粒体和叶绿体的RNA代谢,调节光合作用、呼吸作用、胚胎发生等生理和生长发育过程;响应逆境胁迫 Mainly participating in the RNA metabolism of mitochondrial and chloroplast, regulating photosynthesis, respiration, embryogenesis and other physiological and growth processes; Responsing to adversity stress |
| DEAD-box RHs | 包含Q、Ⅰ、Ⅱ（DEAD）、Ⅲ、Ⅳ、Ⅴ和Ⅵ结构域 Containing Q, Ⅰ, Ⅱ(DEAD), Ⅲ, Ⅳ, Ⅴ and Ⅵ domains | 催化RNA分子二级结构解旋,参与RNA代谢,调节植物各种细胞代谢途径、生长发育;响应低温、干旱等非生物胁迫 Catalyzing the secondary structure of RNA molecules to unwind, participating in RNA metabolism, regulating various cellular metabolic pathways, growth and development; Responding to cold, drought and other abiotic stresses |
| RNA分子伴侣 RNA molecular chaperone | 具有RNA伴侣蛋白活性的RBP,结构特征多样 The structural features of the RBPs with RNA chaperone activity are diverse | 参与非生物胁迫下RNA的折叠反应,提高植物的胁迫耐受性;参与RNA代谢,调节叶绿体相关生物合成等过程 Participating in the folding reaction of RNA under abiotic stresses, thereby improving the stress tolerance of plants; participating in RNA metabolism, regulating the process of chloroplast-related biosynthesis, etc. |
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Table 2
表2
表2植物RNA结合蛋白的分类、物种分布及研究现状
Table 2
| 分类 Classification | 各物种中的数量 Number in each species | 已报道的关键基因 Key genes that have been reported | 参考文献 References |
|---|---|---|---|
| SR蛋白 SR proteins | 拟南芥：19个 水稻：22个 大豆：25个 Arabidopsis: 19 Oryza sativa: 22 Glycine max: 25 | SRp33, SCL26, RSZp23, RSZ33/36/37, RS40/41 | [8-10] |
| GR-RBPs (IV of GRPs) | AtGRP2/7/8; OsGRP1/4/6 | [14,18,21] | |
| RZs (IVb of GRPs) | 拟南芥：3个 水稻：3个 小麦：4个 Arabidopsis: 3 Oryza sativa: 3 Triticum aestivum: 4 | AtRZ-1a/b/c; OsRZ1/2/3; TaRZ1/2/3/4 | [23-26] |
| CSDPs (IVc of GRPs) | 拟南芥：4个 水稻：2个 小麦：4个 Arabidopsis: 4 Oryza sativa: 2 Triticum aestivum: 4 | AtCSP1/2/3/4; OsCSP1/2; WCSP1/2/3/4 | [29-30,33-34] |
| PPR蛋白 PPR proteins | 拟南芥：多于450个 水稻：约477个 Arabidopsis: more than 450 Oryza sativa: about 477 | BLX, RPF5, SOAR1,; AtSEL1,OsPPR676; OsRF5,OsWSL | [37,40-41,45,48,51-52] |
| DEAD-box RHs | 拟南芥：约58个 水稻：约50个 Arabidopsis: about 58 Oryza sativa: about 50 | AtRH50, ZmRH3, TOGR1, RID1 | [62-63, 65-66] |
| RNA分子伴侣 RNA molecular chaperone | AtRH3, SRRP1; CFM4 | [80-81,83] |
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1 富含丝氨酸―精氨酸的RNA结合蛋白（SR蛋白）家族
SR蛋白家族在动植物中都是非常重要的一类RNA结合蛋白,并且在高等真核生物中高度保守。SR蛋白参与基因转录后调控的关键步骤,是前体mRNA组成型和选择性剪接的必需因子。SR蛋白的结构特征包括：N端有1个或2个RNA识别基序（RRM）,C端是富含丝氨酸和精氨酸的二肽（dipeptide rich in serine and arginine,SR）结构域。研究较多的动物SR蛋白大多参与mRNA的剪接过程。与人类的12个SR蛋白相比,植物中SR蛋白的种类和数量较多一些,比如拟南芥和水稻中分别有19和22个SR蛋白,大豆具有25个SR蛋白,但是关于这些植物SR蛋白的研究还很少。目前,植物中的研究表明SR蛋白在植物的生长发育和胁迫响应中起着关键作用。水稻SRp32、SRp33a、SRp33b、SCL26、RSZp23、RSZ36和RSZ37a通过参与mRNA的剪接、输出或翻译过程,调节水稻生长发育的各个方面[8];拟南芥RS40和RS41能够调节自身基因前体mRNA和其他SR基因的选择性剪接[9,10],AtRSZ33也具有类似的剪接模式。此外,AtRSZ33的异位表达会导致植物发育异常,比如加快细胞增殖、改变细胞伸长和分裂极性等[11],这表明植物SR蛋白可能通过mRNA代谢过程而实现对植物生长发育的调节。SR蛋白在RNA代谢中的可变性使其在非生物胁迫下具有较强的胁迫弹性,例如温度胁迫和激素处理均能改变SR蛋白的选择性剪接,如受激素影响的SR1、SR34b和SCL33等蛋白,这表明非生物胁迫和激素可能通过改变SR基因转录本的结构而影响它们的选择性剪接功能、剪接效率和靶向性[12]。
植物SR蛋白往往通过与同源或异源蛋白的相互作用来发挥重要的剪接功能。例如水稻RSp29和SCL26相互作用以增加mRNA的剪接效率;RSZ36和SRp33b相互作用以改变自身和其他SR蛋白的选择性剪接模式[8];RS40和RS41蛋白与含KH结构域的RNA结合蛋白HOS5等相互作用共同调节前体mRNA的加工[10]。目前,对植物SR蛋白的研究仍处于初级阶段,很多与之相关的功能和机制尚未得到解析,高等真核生物的SR蛋白具有较高的序列保守性,这或许是研究植物SR蛋白可有效利用的一种捷径。
2 富含甘氨酸的RNA结合蛋白（GR- RBPs）家族
植物中富含甘氨酸的蛋白质（GRPs）以高含量的甘氨酸为特征（20%—70%）,共分为4类,其中只有第Ⅳ类GRPs（GR-RBPs）具有RNA结合功能,是RNA结合蛋白[13]。GR-RBPs在植物中广泛存在,参与调节植物生长发育和响应非生物胁迫过程。GR-RBPs的结构特征是在C-末端存在富含甘氨酸的区域,N-末端存在RNA识别基序（RRM）或冷休克结构域（CSD）。拟南芥基因组编码十多种GR-RBPs,这些GR-RBPs在生长发育以及各种胁迫反应中发挥重要作用。具有RNA分子伴侣活性的AtGRP7和AtGRP2在水稻中的过表达有助于提高水稻的干旱耐受性和种子产量,并且其转基因植株对生长发育无任何副作用,因此,它们可以作为一种潜在的调节基因来改善干旱胁迫下的水稻生长发育[14]。AtGRP7能增强拟南芥对干旱和高盐的耐受性,它通过非ABA依赖型的方式调节气孔开度以控制水分蒸腾,从而介导植物的非生物胁迫反应[15]。此外,AtGRP7能够通过调节保卫细胞中的mRNA输出而提高植株的低温耐受性[15]。AtGRP7是丁香假单胞菌Ⅲ型效应物HopU1的直接靶标,其和AtGRP8不仅能提高拟南芥的免疫功能[16,17],而且还能作为生物钟调节器共同调节昼夜节律[18]。AtGRP7和AtGRP8在自主开花途径中能够调节植物开花,在atgrp7突变体和AtGRP8反转义植株中开花阻遏蛋白（FLOWERING LOCUS C,FLC）被上调,并由此导致晚花表型。与此相反,过表达AtGRP7会降低FLC转录,从而导致早花[19]。AtGRP2以非ABA依赖型的方式促进盐或冷胁迫下的种子萌发和幼苗生长,提高拟南芥盐和低温耐受性[20]。综上所述,拟南芥的GR-RBPs主要作为RNA分子伴侣在非生物胁迫响应中发挥重要作用,同时其可能通过介导植物激素信号通路而调节植物的生长发育。
水稻和拟南芥的GR-RBPs在功能上具有保守性,它们在植物响应低温过程中主要作为RNA分子伴侣发挥调节作用。3个水稻GR-RBP蛋白（OsGRP1、OsGRP4和OsGRP6）能够弥补冷休克期间低温敏感型大肠杆菌突变体的生长缺陷表型,以及恢复冷胁迫下拟南芥突变体atgrp7的缺陷表型[21]。KIM等[21]研究发现OsGRP4和OsGRP6通过介导mRNA从细胞核向细胞质的运输过程来参与水稻的低温胁迫响应。
2.1 含有CCHC型锌指基序的GR-RBP—RZ家族
在GRPs家族的4类蛋白中,第Ⅳ类GRPs又分为a、b、c和d 4个不同的子类,含有CCHC型锌指基序的GRPs属于Ⅳb亚类（RZs家族）。RZs在N-末端含有RRM结构域,在C-末端有富含甘氨酸的区域且其间散布着CCHC型锌指基序。不同物种中的RZs在结构和功能上具有保守性,参与植物的生长发育和胁迫响应过程。拟南芥基因组编码3个RZ基因,分别被命名为AtRZ-1a、AtRZ-1b和AtRZ-1c。AtRZ-1a在低温胁迫下强烈上调,在干旱和脱落酸胁迫下则与之相反。在低温下AtRZ-1a促进种子萌发和幼苗生长,而其突变体中的上述过程明显受到抑制[22,23]。在盐或干旱胁迫条件下,AtRZ-1a则抑制拟南芥种子萌发和幼苗生长,说明AtRZ-1a的调节方式具有环境特异性[22,23]。AtRZ-1b和AtRZ-1c通过与其他蛋白如SR蛋白相互作用,调节前体mRNA剪接,在植物发育的许多方面发挥重要作用。AtRZ-1b和AtRZ-1c双突变植株表现出多种发育缺陷,如种子萌发和根生长延迟、晚花、少花、植株高度降低、叶尺寸减小以及锯齿状叶等[24]。转录组分析发现许多关键的发育调节基因和激素相关基因的表达水平在atrz-1b artz-1c双突变体中发生改变,这是导致双突变体出现多种异常表型的原因。同时,AtRZ-1b和AtRZ-1c是FLC内含子剪接所必须的,AtRZ-1b和AtRZ-1c促进FLC第一个内含子的高效剪接并抑制FLC转录,从而调节植物开花[24]。
水稻基因组也编码3个RZ基因（OsRZ1— OsRZ3）,其转录水平均受低温胁迫诱导,受干旱或盐胁迫的影响微弱。上述3个基因中只有OsRZ2能够提高拟南芥的低温耐受性,表明其功能存在一定差异[25]。小麦基因组编码4个RZ基因（TaRZ1— TaRZ4）,它们在非生物胁迫下植物的种子萌发、幼苗生长和低温耐受性方面发挥不同功能。盐胁迫下,过表达TaRZ1—TaRZ3的3种转基因拟南芥的种子萌发均被抑制;干旱胁迫下,过表达TaRZ2或TaRZ3的转基因拟南芥种子萌发受到抑制;过表达TaRZ1的转基因拟南芥幼苗生长在低温或盐胁迫下被显著抑制,而过表达TaRZ2则提高了拟南芥的低温耐受性[26]。由此推测不同物种的RZs在参与植物的生长发育以及胁迫反应过程中,存在一定的功能保守性。
2.2 含有冷休克结构域的GR-RBPs—CSDPs家族
CSDPs家族属于GR-RBPs的Ⅳc亚类。冷休克结构域（cold shock domain,CSD）是与RNA、ssDNA和dsDNA结合有关的高度保守的核酸结合结构域,在细菌、动物和植物中广泛存在。细菌冷休克蛋白由单一的CSD组成,在低温下被诱导,并在低温胁迫反应过程中发挥重要作用[27]。植物CSD蛋白的结构特征是在N端含有一个CSD,C端含有一个富含甘氨酸的区域,并在富含甘氨酸的区域中散布着不同数量的CCHC型锌指基序[28]。CSDPs同样在植物发育和胁迫响应中发挥重要作用。拟南芥中存在4种CSD蛋白,分别为AtCSP1、AtCSP2、AtCSP3和AtCSP4。其中,AtCSP2表达量最高,AtCSP3是目前研究最深入的,AtCSP2与AtCSP4的同源性最高,且在某些方面具有功能冗余。AtCSP1在干旱或盐胁迫下植物的种子萌发和生长中发挥作用[29]。AtCSP2通过CBF依赖性途径负调节植物的低温耐受性,其过表达导致植物的低温耐受性降低,此外在低温耐受性和开花时间的调控方面与AtCSP4存在功能冗余。另外,过表达AtCSP2导致植株的形态和生长发育发生改变,如植株矮小,开花延迟和果荚变短[30],AtCSP2还能够通过调节GA和ABA代谢基因的表达来负调控种子萌发[28]。AtCSP3参与植物盐和干旱胁迫耐受性的调控,过表达AtCSP3能够显著提高盐和干旱胁迫下的植株存活率[31]。AtCSP4通过调节与发育相关的基因表达而在长角果发育的晚期阶段起重要作用,过表达AtCSP4导致MADS-box和胚乳发育基因的表达在花和果实发育过程中发生异常改变,转基因植株表现出长角果缩短和种子成熟度低等缺陷表型[32]。
小麦也编码4种CSD蛋白（WCSP1—WCSP4）,其中WCSP1是第一个被发现的CSD蛋白,WCSP1能赋予低温敏感型大肠杆菌较强的耐受性[33]。水稻编码2种CSD蛋白,分别为OsCSP1和OsCSP2,它们表现出核酸结合活性并改善了大肠杆菌csp突变体的低温敏感性[34]。此外,小麦WCSPs和水稻OsCSPs在植物发育中也发挥重要作用[33,34],但仍未见相关的研究报道。目前,关于植物CSD蛋白的研究还很少,虽然在植物中已经鉴定出许多CSD蛋白,但对它们功能的解析程度还不够深入。
3 三角状五肽重复区蛋白（PPR蛋白）家族
PPR蛋白是一种序列特异性RNA结合蛋白,它们以序列特异性方式结合RNA,是目前研究最深入和最重要的一类RNA结合蛋白之一。PPR蛋白家族成员是mRNA转录后靶向细胞器的关键调控者,主要参与线粒体和叶绿体的RNA代谢过程。PPR蛋白几乎涉及植物细胞器RNA代谢的所有方面,包括RNA编辑、RNA剪接、RNA稳定以及翻译过程。PPR蛋白由大约35个氨基酸序列经串联重复组成,并且随后折叠成一对反向平行的α螺旋。PPR蛋白主要被分为2个亚类,P类和PLS类。P类PPR蛋白具有规范的35个氨基酸基序,但有些除了PPR基序外,还会在末端存在额外的结构域,以行使特异性功能;而PLS类的PPR蛋白具有3种不同类型的PPR基序,其长度各不相同：P（35个氨基酸）、L（35—36个氨基酸）和S（约31个氨基酸）。PLS类PPR蛋白一般会包含额外的结构域,根据这些结构域的不同,又分为不同的亚家族,包括PLS、E1、E2、E+和DYW等家族,不同类型的PPR蛋白可能具有不同的功能[35,36]。PPR蛋白主要存在于植物中,在其他真核生物中较少,人类基因组中只有不到10个PPR蛋白。在植物中,拟南芥基因组含有超过450个PPR蛋白,水稻中大约有477个,除这两类外,其他物种如玉米中含量相对较少。PPR蛋白在细胞器RNA加工、CMS植物的育性恢复、光合作用、呼吸作用、胚胎发生、植物发育以及胁迫响应过程中发挥重要作用。目前已经有部分植物PPR蛋白被报道,但仍有大部分PPR蛋白的分子功能未得到解析。下面主要总结了近几年关于拟南芥、水稻和玉米PPR蛋白的功能研究进展。在拟南芥中,靶向线粒体的PPR蛋白BLX[37]、AtGRS1[38]和SLO4[39]参与调节线粒体内含子的剪接,并对植物早期胚胎发育和植株生长发挥重要作用。还有一些靶向线粒体的 PPR蛋白,它们不仅在RNA代谢过程中发挥重要作用,而且在改善细胞质雄性不育和逆境胁迫等方面也具有重要功能。如RPF5蛋白除了参与特定mRNA的成熟过程外,也能够部分恢复细胞质雄性不育系的育性[40]。SOAR1是拟南芥响应非生物胁迫重要的正调控蛋白,过表达SOAR1能显著提高植物的干旱、盐和低温的胁迫耐受性,并且不会影响植物的正常生长和发育。SOAR1还是ABA信号转导的关键负调控因子,通过整合ABA依赖型和非依赖型的信号传导途径来调节植物的应激反应[41,42]。AtSLG1在RNA编辑、植物生长发育、ABA处理和干旱胁迫反应中发挥重要作用 [43]。AtPGN在植物响应逆境胁迫过程中发挥重要作用。pgn突变体表现出对坏死性真菌病原体的易感性以及对脱落酸、葡萄糖和盐胁迫的超敏感性。基因功能研究揭示PGN在胁迫反应过程中通过调节线粒体的活性氧动态平衡来发挥作用[44]。靶向叶绿体的拟南芥PPR蛋白,它们除了在叶绿体内含子剪接过程中起至关重要的作用外,在生长发育以及光合作用中也发挥重要作用。如AtSEL1参与叶绿体发育所需的质体基因的表达调控,在sel1突变体中,光合作用的相关蛋白减少且叶绿体发育受损[45];AtPPR2在胚胎发育的第一次有丝分裂和细胞增殖过程中发挥重要作用[46];此外还有叶绿体发育及光合作用所必须的SVR7蛋白,其基因突变体（svr7）显示叶绿体ATP合酶亚基的积累受到影响。综上所述,拟南芥PPR蛋白在细胞器mRNA代谢、植物生长发育以及胁迫反应等方面发挥关键作用[47]。
在水稻中,许多PPR蛋白除了具有基本的RNA编辑功能外,还参与植物发育的各个方面,如花粉发育、幼苗生长和叶绿体发育,并且在改善细胞质雄性不育和非生物胁迫等方面也发挥重要作用。如OsPPR676蛋白是水稻生长和花粉发育所必需的,OsPPR676对于质体atpB亚基的产生是必不可少的,其通过影响ATP合酶的活性在脂肪酸、碳水化合物以及其他有机物的生物合成中发挥关键作用[48]。OsPPR6参与叶绿体中与光合作用相关的基因转录物的剪接并调节早期叶绿体的生物合成,是水稻叶绿体生物合成所必需的调节因子[49]。OsPGL1通过参与线粒体和叶绿体中特定位点的RNA编辑而调节相关的生物过程[50]。OsRF5通过与富含甘氨酸的蛋白质GRP162相互作用而恢复红莲型细胞质雄性不育系的育性[51]。OsWSL[52]和OsWSL4[53]是叶绿体重要的剪接因子,影响水稻早期叶片发育,且OsWSL能提高植物对ABA、盐和糖胁迫的耐受性。OsV4在低温胁迫下早期的叶绿体发育中起重要作用[54]。
近期发现玉米PPR蛋白在基因功能上存在一定的相似性。玉米PPR蛋白的种类较少,且大多靶向线粒体。如靶向线粒体的EMP10[55]、EMP11[56]、DEK10[57]和DEK35[58]蛋白对线粒体内mRNA的不同位点进行RNA编辑和剪接,是维持线粒体功能和胚胎、胚乳发育不可或缺的一类PPR蛋白。另外,PPR78[59]和PPR2263[60]对线粒体mRNA进行编辑和剪接,对线粒体发挥功能和种子发育具有重要作用。近几年,尽管已经鉴定出许多植物PPR蛋白,但由于植物中PPR蛋白本身基数较大的缘故,还有很多发挥重要功能的PPR蛋白未被研究。此外,目前关于PPR蛋白作用的详细地分子机制还不清楚,需要进一步深入探究。
4 DEAD-box RNA解旋酶（DEAD-box RHs）家族
RNA解旋酶（RH）是能够改变RNA结构的酶。RHs分为6个超家族（SF1—SF6）,其中超家族Ⅱ（SF2）是最大的解旋酶家族,主要由DEAD-box RHs组成, DEAD-box RHs具有RNA结合活性,是RNA结合蛋白。DEAD-box RHs可以催化RNA分子中二级结构的解旋,从而影响有机体中RNA代谢过程,它们在调节各种细胞代谢途径中发挥重要作用。DEAD-box RNA解旋酶由Q、Ⅰ、Ⅱ（DEAD）、Ⅲ、Ⅳ、Ⅴ和Ⅵ结构域组成,这些结构域具有不同的功能,有的具有解旋功能,有的具有RNA结合活性,有的具有其他特异性功能。在真核生物中,植物编码的DEAD-box RHs更多,拟南芥编码约58个,水稻约50个,人类基因组中约有36个。目前植物中的DEAD-box RHs研究主要集中在拟南芥和水稻上,DEAD-box RHs除了在RNA代谢方面发挥重要作用外,在植物生长发育和非生物胁迫中也行使重要功能。4.1 DEAD-box RHs在RNA代谢中起着至关重要的作用
大多数DEAD-box RHs在RNA代谢过程中发挥作用,这些DEAD-box RHs不仅是细胞核RNA代谢中必不可少的调节因子,有些也参与线粒体和叶绿体的RNA代谢过程。如AtRH9参与线粒体mRNA中Ⅱ组内含子的剪接[61];AtRH50参与叶绿体23S和4.5S rRNA的加工[62];ZmRH3参与rRNA的生物合成以及叶绿体中mRNA的剪接[63];RCF1[64]和RID1[65]是拟南芥前体mRNA必要的剪接因子;水稻TOGR1[66]在高温下能维持rRNA的稳定性。综上所述,DEAD-box RHs在RNA代谢中起着重要作用。4.2 DEAD-box RHs在植物生长发育和非生物胁迫反应中具有不同作用
许多DEAD-box RHs在植物生长发育过程中也具有重要作用。如RID1在配子体发育过程中发挥重要作用[65];AtRH57参与植物种子萌发和幼苗生长过程中葡萄糖和脱落酸信号交联的糖应答过程[67];拟南芥AtRH36[68]和水稻OsRH36[69]参与雌配子的有丝分裂过程,它们在功能上具有保守性;OsRH2和OsRH34参与调控水稻株高、花粉和种子发育[70],综上所述,DEAD-box RHs在植物生长发育的多个过程中发挥重要的调节作用。DEAD-box RHs在非生物胁迫中的潜在作用也日益被发现。如RCF1参与植物低温感应基因的调控并增加植株的低温耐受性[64];AtRH7参与植物发育和拟南芥的低温耐受性[71];过表达TOGR1促进了水稻在高温胁迫下的生长[66];靶向线粒体的拟南芥AtRH9和AtRH25的过表达抑制了高盐浓度条件下的种子发芽,其中 AtRH25的过表达也能增强植株的低温耐受性[72];RH8与PP2CA相互作用,通过ABA依赖型信号传导来调节干旱胁迫反应[73];叶绿体靶向的BrRH22在高盐和干旱胁迫下促进拟南芥种子发芽和植株生长[74]。以上结果表明DEAD-box RHs在非生物胁迫反应中扮演关键作用。尽管DEAD-box RHs在植物中已取得较大进展,但仍有很多DEAD-box RHs未被鉴定,此外关于DEAD-box RHs参与的信号转导调控网络还不清楚,因此,未来还要进一步确定它的RNA靶标及其与其他调节信号的交联机制。
5 RNA分子伴侣
同蛋白质一样,RNA分子需要正确地折叠成相应的结构才能发挥正常功能,然而RNA分子由于其固有的动力学和热力学折叠问题而容易错误折叠成非功能性结构。RNA分子伴侣是非特异性的RNA结合蛋白,通过对错误折叠的RNA进行结构重排来促进RNA的正确折叠,以保证RNA代谢过程的顺利进行[75]。RNA分子伴侣在生物体如细菌、病毒、动物和植物的生长和发育中起着关键作用,其中对RNA分子伴侣在植物中的研究远远落后于其他3类生物。许多富含甘氨酸的RNA结合蛋白（GR-RBPs）以及DEAD-box RNA解旋酶（DEAD-box RHs）家族成员具有RNA分子伴侣活性,它们在植物生长发育和胁迫反应中发挥重要作用,并且有些已被确定在逆境胁迫下作为RNA分子伴侣起作用。具有RNA分子伴侣活性的RNA结合蛋白参与植物胁迫响应。当细胞暴露于低温时,RNA分子伴侣的作用更加突出,错误折叠的RNA分子在低温下变得过度稳定,在缺少RNA分子伴侣的帮助下不能呈现天然构象。研究证明GR-RBPs的某些家族成员在低温反应期间表现出RNA分子伴侣活性,如拟南芥的AtGRP2 [20]和AtGRP7[21];水稻的OsGRP1、OsGRP4和OsGRP6 [21]等蛋白。另外,在GR-RBPs家族中,含有CCHC型锌指基序的GR-RBPs（RZs）成员也被发现具有RNA分子伴侣活性,如拟南芥AtRZ-1a[76]、水稻OsRZ2[25]和小麦TaRZ2[77]等蛋白。GR-RBPs家族的CSDP1[78]在提高拟南芥低温耐受性方面也起RNA分子伴侣作用。以上结果表明具有RNA分子伴侣活性的GR-RBPs家族成员在植物胁迫反应中发挥重要作用。在细胞响应低温过程中,GR-RBPs的结构域序列和总体折叠方式对于GR-RBPs的RNA分子伴侣活性至关重要[79]。DEAD-box RHs家族的某些成员在植物胁迫反应中也具有RNA分子伴侣活性,如低温胁迫下的AtRH25蛋白[72];另外由具有RNA分子伴侣活性的拟南芥AtRH3 所调节的内含子剪接对于叶绿体功能、植物的生长和胁迫反应是至关重要的[80]。
RNA分子伴侣在植物的生长和发育中起着关键作用,尤其是一些靶向叶绿体的RNA分子伴侣。如靶向叶绿体的SRRP1影响拟南芥叶绿体转录物的剪接和加工[81];靶向叶绿体的SDP影响rRNA加工、叶绿体生物合成及光合作用[82],这些对拟南芥的正常生长是至关重要的;还有CFM4通过参与叶绿体rRNA的加工在拟南芥生长和应激反应中发挥作用[83]。综上所述,在植物生长和发育过程中,许多具有RNA分子伴侣活性的RNA结合蛋白在RNA代谢的调节过程中发挥重要作用。目前,已有许多证据表明RNA分子伴侣在植物生长发育以及胁迫反应中发挥重要作用,但关于RNA分子伴侣的研究还不够深入,接下来不仅要继续挖掘更多具有RNA分子伴侣活性的RBPs,还要研究RNA分子伴侣如何识别底物RNA以及它们如何与其他蛋白协作共同参与转录后的RNA代谢过程,从而调节植物生长发育并响应环境胁迫。
6 结论与展望
植物中的RBPs作为重要的转录后调控因子,在RNA代谢、植物生长发育以及胁迫反应过程中发挥重要作用。在本文所论述的五类植物RBP家族中,SR蛋白主要作为重要的选择性剪接因子在植物的生长发育和胁迫响应中起着关键作用;GR-RBPs家族的成员普遍具有功能多样性,其可能通过参与介导植物激素信号通路调节植物生长发育的多个方面,并主要作为RNA分子伴侣在多种胁迫反应中发挥重要作用;PPR蛋白主要参与线粒体和叶绿体的RNA代谢,调节植物生长发育以及胁迫反应过程;DEAD-box RHs可作为细胞核和细胞器重要的RNA剪接因子,并在植物生长发育以及非生物胁迫反应中发挥多种功能;RNA分子伴侣作为非特异性的RNA结合蛋白,通过参与RNA折叠反应保证RNA代谢过程的顺利进行,此外,许多RBPs具有RNA分子伴侣活性,它们作为RNA分子伴侣调节植物的生长发育以及胁迫反应。总的来说,上述五类RNA结合蛋白在植物中的作用集中体现在RNA代谢和RNA折叠反应方面,并通过这两个方面的作用,影响相关基因的表达,从而调节胚胎发育、幼苗生长等生长发育以及响应干旱、低温等逆境胁迫。近年来,关于RBPs在植物生长发育和胁迫反应中的研究正在迅速增加,目前,关于SR蛋白以及RNA分子伴侣的研究还很少,对于相对研究较多的GR-RBPs和PPR蛋白来说,应进一步深入确定它们的RNA靶标及其参与的信号转导调控网络。很多RBPs在高等真核生物中具有较强的保守性,因此,可以利用植物中的某些蛋白与动物RNA结合蛋白的同源性去寻找植物中具有潜在功能的RBPs。此外,随着近年来转录组测序技术的发展使我们能够获得植物在某一生理状态下的转录组信息,从而通过对RNA结合蛋白突变体或逆境胁迫中的转录组进行组学分析,预测RNA结合蛋白的潜在功能和可能参与的信号调控网络。未来关于RBPs的研究应该更加系统和全面化,除了继续挖掘更多的具有重要功能的RBPs外,还应关注RBPs作用的RNA靶标,探究RBPs识别底物RNA的生化机制,以及RBPs与其他调控因子交联而共同调节植物生命活动的信号调控网络。
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被引期刊影响因子
DOI:10.1016/j.molcel.2014.04.033URLPMID:24856220 [本文引用: 1]
The proper processing, export, localization, translation, and degradation of mRNAs are necessary for regulation of gene expression. These processes are controlled by mRNA-specific regulatory proteins, noncoding RNAs, and core machineries common to most mRNAs. These factors bind the mRNA in large complexes known as messenger ribonucleoprotein particles (mRNPs). Herein, we review the components of mRNPs, how they assemble and rearrange, and how mRNP composition differentially affects mRNA biogenesis, function, and degradation. We also describe how properties of the mRNP “interactome” lead to emergent principles affecting the control of gene expression.
DOI:10.1146/annurev-biochem-080111-092106URLPMID:25784054 [本文引用: 1]
Throughout their lifetimes, messenger RNAs (mRNAs) associate with proteins to form ribonucleoproteins (mRNPs). Since the discovery of the first mRNP component more than 40 years ago, what is known as the mRNA interactome now comprises >1,000 proteins. These proteins bind mRNAs in myriad ways with varying affinities and stoichiometries, with many assembling onto nascent RNAs in a highly ordered process during transcription and precursor mRNA (pre-mRNA) processing. The nonrandom distribution of major mRNP proteins observed in transcriptome-wide studies leads us to propose that mRNPs are organized into three major domains loosely corresponding to 5′ untranslated regions (UTRs), open reading frames, and 3′ UTRs. Moving from the nucleus to the cytoplasm, mRNPs undergo extensive remodeling as they are first acted upon by the nuclear pore complex and then by the ribosome. When not being actively translated, cytoplasmic mRNPs can assemble into large multi-mRNP assemblies or be permanently disassembled and degrade...
DOI:10.1093/nar/30.3.623URLPMID:100298 [本文引用: 1]
Regulation of gene expression at the post-transcriptional level is mainly achieved by proteins containing well-defined sequence motifs involved in RNA binding. The most widely spread motifs are the RNA recognition motif (RRM) and the K homology (KH) domain. In this article, we survey the complete Arabidopsis thaliana genome for proteins containing RRM and KH RNA-binding domains. The Arabidopsis genome encodes 196 RRM-containing proteins, a more complex set than found in Caenorhabditis elegans and Drosophila melanogaster. In addition, the Arabidopsis genome contains 26 KH domain proteins. Most of the Arabidopsis RRM-containing proteins can be classified into structural and/or functional groups, based on similarity with either known metazoan or Arabidopsis proteins. Approximately 50% of Arabidopsis RRM-containing proteins do not have obvious homologues in metazoa, and for most of those that are predicted to be orthologues of metazoan proteins, no experimental data exist to confirm this. Additionally, the function of most Arabidopsis RRM proteins and of all KH proteins is unknown. Based on the data presented here, it is evident that among all eukaryotes, only those RNA-binding proteins that are involved in the most essential processes of post-transcriptional gene regulation are preserved in structure and, most probably, in function. However, the higher complexity of RNA-binding proteins in Arabidopsis, as evident in groups of SR splicing factors and poly(A)-binding proteins, may account for the observed differences in mRNA maturation between plants and metazoa. This survey provides a first systematic analysis of plant RNA-binding proteins, which may serve as a basis for functional characterisation of this important protein group in plants.
DOI:10.14348/molcells.2016.2359URLPMID:4794599 [本文引用: 1]
Posttranscriptional regulation of RNA metabolism, including RNA processing, intron splicing, editing, RNA export, and decay, is increasingly regarded as an essential step for fine-tuning the regulation of gene expression in eukaryotes. RNA-binding proteins (RBPs) are central regulatory factors controlling posttranscriptional RNA metabolism during plant growth, development, and stress responses. Although functional roles of diverse RBPs in living organisms have been determined during the last decades, our understanding of the functional roles of RBPs in plants is lagging far behind our understanding of those in other organisms, including animals, bacteria, and viruses. However, recent functional analysis of multiple RBP family members involved in plant RNA metabolism and elucidation of the mechanistic roles of RBPs shed light on the cellular roles of diverse RBPs in growth, development, and stress responses of plants. In this review, we will discuss recent studies demonstrating the emerging roles of multiple RBP family members that play essential roles in RNA metabolism during plant growth, development, and stress responses.
DOI:10.1016/j.febslet.2008.03.004URLPMID:2858862 [本文引用: 1]
RNAs in cells are associated with RNA-binding proteins (RBPs) to form ribonucleoprotein (RNP) complexes. The RBPs influence the structure and interactions of the RNAs and play critical roles in their biogenesis, stability, function, transport and cellular localization. Eukaryotic cells encode a large number of RBPs (thousands in vertebrates), each of which has unique RNA-binding activity and protein rotein interaction characteristics. The remarkable diversity of RBPs, which appears to have increased during evolution in parallel to the increase in the number of introns, allows eukaryotic cells to utilize them in an enormous array of combinations giving rise to a unique RNP for each RNA. In this short review, we focus on the RBPs that interact with pre-mRNAs and mRNAs and discuss their roles in the regulation of post-transcriptional gene expression.
DOI:10.1186/1471-2407-14-322URLPMID:24884756 [本文引用: 1]
Background RNA binding proteins (RBPs) play a fundamental role in posttranscriptional control of gene expression. Different RBPs have oncogenic or tumor-suppressive functions on human cancers. RNPC1 belongs to the RNA recognition motif (RRM) family of RBPs, which could regulate expression of diverse targets by mRNA stability in human cancer cells. Several studies reported that RNPC1 played an important role in cancer, mostly acting as an oncogene or up-regulating in tumors. However, its role in human breast cancer remains unclear. Methods In the present study, we investigated the functional and mechanistic roles of RNPC1 in attenuating invasive signal including reverse epithelial-mesenchymal transition (EMT) to inhibit breast cancer cells aggressiveness in vitro. Moreover, RNPC1 suppress tumorigenicity in vivo. Further, we studied the expression of RNPC1 in breast cancer tissue and adjacent normal breast tissue by quantitative RT-PCR (qRT-PCR) and Western blot. Results We observed that RNPC1 expression was silenced in breast cancer cell lines compared to breast epithelial cells. More important, RNPC1 was frequently silenced in breast cancer tissue compared to adjacent normal breast tissue. Low RNPC1 mRNA expression was associated with higher clinical stages and mutp53, while low level of RNPC1 protein was associated with higher lymph node metastasis, mutp53 and lower progesterone receptor (PR). Functional assays showed ectopic expression of RNPC1 could inhibit breast tumor cell proliferation in vivo and in vitro through inducing cell cycle arrest, and further suppress tumor cell migration and invasion partly through repressing mutant p53 (mutp53) induced EMT. Conclusions Overall, our findings indicated that RNPC1 had a potential function to play a tumor-suppressor role which may be a potential marker in the therapeutic and prognostic of breast cancer.
DOI:10.1007/s00018-015-1943-xURLPMID:4565867 [本文引用: 1]
The loss of synapses is a central event in neurodegenerative diseases. Synaptic proteins are often associated with disease neuropathology, but their role in synaptic loss is not fully understood. Of the many processes involved in sustaining the integrity of synapses, local protein translation can directly impact synaptic formation, communication, and maintenance. RNA-binding proteins and their association with RNA granules serve to regulate mRNA transportation and translation at synapses and in turn regulate the synapse. Genetic mutations in RNA-binding proteins FUS and TDP-43 have been linked with causing neurodegenerative diseases: amyotrophic lateral sclerosis and frontotemporal dementia. The observation that mutations in FUS and TDP-43 coincide with changes in RNA granules provides evidence that dysfunction of RNA metabolism may underlie the mechanism of synaptic loss in these diseases. However, we do not know how mutations in RNA-binding proteins would affect RNA granule dynamics and local translation, or if these alterations would cause neurodegeneration. Further investigation into this area will lead to important insights into how disruption of RNA metabolism and local translation at synapses can cause neurodegenerative diseases.
DOI:10.1105/tpc.105.037069URL [本文引用: 2]
Ser/Arg-rich (SR) proteins play important roles in the constitutive and alternative splicing of pre-mRNA. We isolated 20 rice (Oryza sativa) genes encoding SR proteins, of which six contain plant-specific characteristics. To determine whether SR proteins modulate splicing efficiency and alternative splicing of pre-mRNA in rice, we used transient assays in rice protoplasts by cotransformation of SR protein genes with the rice$Waxy^{b}(Wx^{b})$- -glucuronidase fusion gene. The results showed that plant-specific RSp29 and RSZp23, an SR protein homologous to human 9G8, enhanced splicing and altered the alternative 5 splice sites of$Wx^{b}$intron 1. The resulting splicing pattern was unique to each SR protein; RSp29 stimulated splicing at the distal site, and RSZp23 enhanced splicing at the proximal site. Results of domain-swapping experiments between plant-specific RSp29 and SCL26, which is a homolog of human SC35, showed the importance of RNA recognition motif 1 and the Arg/Ser-rich (RS) domain for the enhancement of splicing efficiencies. Overexpression of plant-specific RSZ36 and SRp33b, a homolog of human ASF/SF2, in transgenic rice changed the alternative splicing patterns of their own pre-mRNAs and those of other SR proteins. These results show that SR proteins play important roles in constitutive and alternative splicing of rice pre-mRNA.
DOI:10.1093/nar/gkr932URLPMID:3315328 [本文引用: 1]
Alternative splicing (AS) coupled to nonsense-mediated decay (NMD) is a post-transcriptional mechanism for regulating gene expression. We have used a high-resolution AS RT–PCR panel to identify endogenous AS isoforms which increase in abundance when NMD is impaired in the Arabidopsis NMD factor mutants, upf1-5 and upf3-1. Of 270 AS genes (950 transcripts) on the panel, 102 transcripts from 97 genes (32%) were identified as NMD targets. Extrapolating from these data around 13% of intron-containing genes in the Arabidopsis genome are potentially regulated by AS/NMD. This cohort of naturally occurring NMD-sensitive AS transcripts also allowed the analysis of the signals for NMD in plants. We show the importance of AS in introns in 5′ or 3′UTRs in modulating NMD-sensitivity of mRNA transcripts. In particular, we identified upstream open reading frames overlapping the main start codon as a new trigger for NMD in plants and determined that NMD is induced if 3′-UTRs were >35065nt. Unexpectedly, although many intron retention transcripts possess NMD features, they are not sensitive to NMD. Finally, we have shown that AS/NMD regulates the abundance of transcripts of many genes important for plant development and adaptation including transcription factors, RNA processing factors and stress response genes.
DOI:10.1371/journal.pgen.1003875URLPMID:24146632 [本文引用: 2]
Eukaryotic genomes encode hundreds of RNA-binding proteins, yet the functions of most of these proteins are unknown. In a genetic study of stress signal transduction in Arabidopsis, we identified a K homology (KH)-domain RNA-binding protein, HOS5 (High Osmotic Stress Gene Expression 5), as required for stress gene regulation and stress tolerance. HOS5 was found to interact with FIERY2/RNA polymerase II (RNAP II) carboxyl terminal domain (CTD) phosphatase-like 1 (FRY2/CPL1) both in vitro and in vivo. This interaction is mediated by the first double-stranded RNA-binding domain of FRY2/CPL1 and the KH domains of HOS5. Interestingly, both HOS5 and FRY2/CPL1 also interact with two novel serine-arginine (SR)-rich splicing factors, RS40 and RS41, in nuclear speckles. Importantly, FRY2/CPL1 is required for the recruitment of HOS5. In fry2 mutants, HOS5 failed to be localized in nuclear speckles but was found mainly in the nucleoplasm. hos5 mutants were impaired in mRNA export and accumulated a significant amount of mRNA in the nuclei, particularly under salt stress conditions. Arabidopsis mutants of all these genes exhibit similar stress-sensitive phenotypes. RNA-seq analyses of these mutants detected significant intron retention in many stress-related genes under salt stress but not under normal conditions. Our study not only identified several novel regulators of pre-mRNA processing as important for plant stress response but also suggested that, in addition to RNAP II CTD that is a well-recognized platform for the recruitment of mRNA processing factors, FRY2/CPL1 may also recruit specific factors to regulate the co-transcriptional processing of certain transcripts to deal with environmental challenges.
DOI:10.1091/mbc.E03-02-0109URLPMID:12972547 [本文引用: 1]
Abstract Splicing provides an additional level in the regulation of gene expression and contributes to proteome diversity. Herein, we report the functional characterization of a recently described plant-specific protein, atRSZ33, which has characteristic features of a serine/arginine-rich protein and the ability to interact with other splicing factors, implying that this protein might be involved in constitutive and/or alternative splicing. Overexpression of atRSZ33 leads to alteration of splicing patterns of atSRp30 and atSRp34/SR1, indicating that atRSZ33 is indeed a splicing factor. Moreover, atRSZ33 is a regulator of its own expression, as splicing of its pre-mRNA is changed in transgenic plants. Investigations by promoter-beta-glucuronidase (GUS) fusion and in situ hybridization revealed that atRSZ33 is expressed during embryogenesis and early stages of seedling formation, as well as in flower and root development. Ectopic expression of atRSZ33 caused pleiotropic changes in plant development resulting in increased cell expansion and changed polarization of cell elongation and division. In addition, changes in activity of an auxin-responsive promoter suggest that auxin signaling is disturbed in these transgenic plants.
DOI:10.1111/j.1365-313X.2006.03020.xURLPMID:17319848 [本文引用: 1]
Precursor mRNAs with introns can undergo alternative splicing (AS) to produce structurally and functionally different proteins from the same gene. Here, we show that the pre-mRNAs of Arabidopsis genes that encode serine/arginine-rich (SR) proteins, a conserved family of splicing regulators in eukaryotes, are extensively alternatively spliced. Remarkably about 95 transcripts are produced from only 15 genes, thereby increasing the complexity of the SR gene family transcriptome by six-fold. The AS of some SR genes is controlled in a developmental and tissue-specific manner. Interestingly, among the various hormones and abiotic stresses tested, temperature stress (cold and heat) dramatically altered the AS of pre-mRNAs of several SR genes, whereas hormones altered the splicing of only three SR genes. These results indicate that abiotic stresses regulate the AS of the pre-mRNAs of SR genes to produce different isoforms of SR proteins that are likely to have altered function(s) in pre-mRNA splicing. Sequence analysis of splice variants revealed that predicted proteins from a majority of these variants either lack one or more modular domains or contain truncated domains. Because of the modular nature of the various domains in SR proteins, the proteins produced from splice variants are likely to have distinct functions. Together our results indicate that Arabidopsis SR genes generate surprisingly large transcriptome complexity, which is altered by stresses and hormones.
DOI:10.1007/s00438-015-1144-1URLPMID:26589419 [本文引用: 1]
Abstract This study focused on the identification and phylogenetic analysis of glycine-rich RNA binding proteins that contain an RNA recognition motif (RRM)-type RNA binding domain in addition to a region with contiguous glycine residues in representative plant species. In higher plants, glycine-rich proteins with an RRM have met considerable interest as they are responsive to environmental cues and play a role in cold tolerance, pathogen defense, flowering time control, and circadian timekeeping. To identify such RRM containing proteins in plant genomes we developed an RRM profile based on the known glycine-rich RRM containing proteins in the reference plant Arabidopsis thaliana. The application of this remodeled RRM profile that omitted sequences from non-plant species reduced the noise when searching plant genomes for RRM proteins compared to a search performed with the known RRM_1 profile. Furthermore, we developed an island scoring function to identify regions with contiguous glycine residues, using a sliding window approach. This approach tags regions in a protein sequence with a high content of the same amino acid, and repetitive structures score higher. This definition of repetitive structures in a fixed sequence length provided a new glance for characterizing patterns which cannot be easily described as regular expressions. By combining the profile-based domain search for well-conserved regions (the RRM) with a scoring technique for regions with repetitive residues we identified groups of proteins related to the A. thaliana glycine-rich RNA binding proteins in eight plant species.
DOI:10.1016/j.plantsci.2013.10.006URLPMID:24268168 [本文引用: 1]
Although posttranscriptional regulation of RNA metabolism is increasingly recognized as a key regulatory process in plant response to environmental stresses, reports demonstrating the importance of RNA metabolism control in crop improvement under adverse environmental stresses are severely limited. To investigate the potential use of RNA-binding proteins (RBPs) in developing stress-tolerant transgenic crops, we generated transgenic rice plants (Oryza sativa) that express Arabidopsis thaliana glycine-rich RBP (AtGRP) 2 or 7, which have been determined to harbor RNA chaperone activity and confer stress tolerance in Arabidopsis, and analyzed the response of the transgenic rice plants to abiotic stresses. AtGRP2- or AtGRP7-expressing transgenic rice plants displayed similar phenotypes comparable with the wild-type plants under high salt or cold stress conditions. By contrast, AtGRP2- or AtGRP7-expressing transgenic rice plants showed much higher recovery rates and grain yields compared with the wild-type plants under drought stress conditions. The higher grain yield of the transgenic rice plants was due to the increases in filled grain numbers per panicle. Collectively, the present results show the importance of posttranscriptional regulation of RNA metabolism in plant response to environmental stress and suggest that GRPs can be utilized to improve the yield potential of crops under stress conditions.
DOI:10.1111/j.1365-313X.2008.03518.xURLPMID:18410480 [本文引用: 2]
Despite the fact that glycine-rich RNA-binding proteins (GRPs) have been implicated in the responses of plants to environmental stresses, their physiological functions and mechanisms of action in stress responses remain largely unknown. Here, we assessed the functional roles of GRP7, one of the eight GRP family members in Arabidopsis thaliana , on seed germination, seedling growth, and stress tolerance under high salinity, drought, or cold stress conditions. The transgenic Arabidopsis plants overexpressing GRP7 under the control of the cauliflower mosaic virus 35S promoter displayed retarded germination and poorer seedling growth compared with the wild-type plants and T-DNA insertional mutant lines under high salinity or dehydration stress conditions. By contrast, GRP7 overexpression conferred freezing tolerance in Arabidopsis plants. GRP7 is expressed abundantly in the guard cells, and has been shown to influence the opening and closing of the stomata, in accordance with the prevailing stress conditions. GRP7 is localized to both the nucleus and the cytoplasm, and is involved in the export of mRNAs from the nucleus to the cytoplasm under cold stress conditions. Collectively, these results provide compelling evidence that GRP7 affects the growth and stress tolerance of Arabidopsis plants under high salt and dehydration stress conditions, and also confers freezing tolerance, particularly via the regulation of stomatal opening and closing in the guard cells.
DOI:10.1038/emboj.2013.15URLPMID:23395902 [本文引用: 1]
Pathogens target important components of host immunity to cause disease. The Pseudomonas syringae type III-secreted effector HopU1 is a mono-ADP-ribosyltransferase required for full virulence on Arabidopsis thaliana. HopU1 targets several RNA-binding proteins including GRP7, whose role in immunity is still unclear. Here, we show that GRP7 associates with translational components, as well as with the pattern recognition receptors FLS2 and EFR. Moreover, GRP7 binds specifically FLS2 and EFR transcripts in vivo through its RNA recognition motif. HopU1 does not affect the protein protein associations between GRP7, FLS2 and translational components. Instead, HopU1 blocks the interaction between GRP7 and FLS2 and EFR transcripts in vivo. This inhibition correlates with reduced FLS2 protein levels upon Pseudomonas infection in a HopU1-dependent manner. Our results reveal a novel virulence strategy used by a microbial effector to interfere with host immunity.
DOI:10.1038/nature05737URL [本文引用: 1]
DOI:10.1371/journal.pcbi.1002986URLPMID:3610657 [本文引用: 1]
The circadian clock controls many physiological processes in higher plants and causes a large fraction of the genome to be expressed with a 24h rhythm. The transcripts encoding the RNA-binding proteins AtGRP7 (Arabidopsis thaliana Glycine Rich Protein 7) and AtGRP8 oscillate with evening peaks. The circadian clock components CCA1 and LHY negatively affect AtGRP7 expression at the level of transcription. AtGRP7 and AtGRP8, in turn, negatively auto-regulate and reciprocally cross-regulate post-transcriptionally: high protein levels promote the generation of an alternative splice form that is rapidly degraded. This clock-regulated feedback loop has been proposed to act as a molecular slave oscillator in clock output. While mathematical models describing the circadian core oscillator in Arabidopsis thaliana were introduced recently, we propose here the first model of a circadian slave oscillator. We define the slave oscillator in terms of ordinary differential equations and identify the model's parameters by an optimization procedure based on experimental results. The model successfully reproduces the pertinent experimental findings such as waveforms, phases, and half-lives of the time-dependent concentrations. Furthermore, we obtain insights into possible mechanisms underlying the observed experimental dynamics: the negative auto-regulation and reciprocal cross-regulation via alternative splicing could be responsible for the sharply peaking waveforms of the AtGRP7 and AtGRP8 mRNA. Moreover, our results suggest that the AtGRP8 transcript oscillations are subordinated to those of AtGRP7 due to a higher impact of AtGRP7 protein on alternative splicing of its own and of the AtGRP8 pre-mRNA compared to the impact of AtGRP8 protein. Importantly, a bifurcation analysis provides theoretical evidence that the slave oscillator could be a toggle switch, arising from the reciprocal cross-regulation at the post-transcriptional level. In view of this, transcriptional repression of AtGRP7 and AtGRP8 by LHY and CCA1 induces oscillations of the toggle switch, leading to the observed high-amplitude oscillations of AtGRP7 mRNA.
DOI:10.1111/j.1365-313X.2008.03591.xURLPMID:18573194 [本文引用: 1]
The RNA binding protein At GRP7 is part of a circadian slave oscillator in Arabidopsis thaliana that negatively autoregulates its own mRNA, and affects the levels of other transcripts. Here, we identify a novel role for AtGRP7 as a flowering-time gene. An atgrp7-1 T-DNA mutant flowers later than wild-type plants under both long and short days, and independent RNA interference lines with reduced levels of At GRP7, and the closely related At GRP8 protein, are also late flowering, particularly in short photoperiods. Consistent with the retention of a photoperiodic response, the transcript encoding the key photoperiodic regulator CONSTANS oscillates with a similar pattern in atgrp7-1 and wild-type plants. In both the RNAi lines and in the atgrp7-1 mutant transcript levels for the floral repressor FLC are elevated. Conversely, in transgenic plants ectopically overexpressing At GRP7, the transition to flowering is accelerated mainly in short days, with a concomitant reduction in FLC abundance. The late-flowering phenotype of the RNAi lines is suppressed by introducing the flc-3 loss-of-function mutation, suggesting that At GRP7 promotes floral transition, at least partly by downregulating FLC . Furthermore, vernalization overrides the late-flowering phenotype. Retention of both the photoperiodic response and vernalization response are features of autonomous pathway mutants, suggesting that At GRP7 is a novel member of the autonomous pathway.
DOI:10.1111/j.1365-313X.2007.03057.xURLPMID:17376161 [本文引用: 2]
Although glycine-rich RNA-binding protein 2 (GRP2) has been implicated in plant responses to environmental stresses, the function and importance of GRP2 in stress responses are largely unknown. Here, we examined the functional roles of GRP2 in Arabidopsis thaliana under high-salinity, cold or osmotic stress. GRP2 affects seed germination of Arabidopsis plants under salt stress, but does not influence seed germination and seedling growth of Arabidopsis plants under osmotic stress. GRP2 accelerates seed germination and seedling growth in Arabidopsis plants under cold stress, and contributes to enhancement of cold and freezing tolerance in Arabidopsis plants. No differences in germination between the wild-type and transgenic plants were observed following addition of abscisic acid (ABA) or glucose, implying that GRP2 affects germination through an ABA-independent pathway. GRP2 complements the cold sensitivity of an Escherichia coli BX04 mutant and exhibits transcription anti-termination activity, suggesting that it has an RNA chaperone activity during the cold adaptation process. Mitochondrial respiration and catalase and peroxidase activities were affected by expression of mitochondrial-localized GRP2 in Arabidopsis plants under cold stress. Proteome analysis revealed that expression of several mitochondrial-encoded genes was modulated by GRP2 under cold stress. These results provide new evidence indicating that GRP2 plays important roles in seed germination, seedling growth and freezing tolerance of Arabidopsis under stress conditions, and that GRP2 exerts its function by modulating the expression and activity of various classes of genes.
URL [本文引用: 4]
DOI:10.1111/j.1365-313X.2005.02420.xURLPMID:15941401 [本文引用: 2]
Summary Glycine-rich RNA-binding proteins (GR-RBPs) have been implicated to play roles in post-transcriptional regulation of gene expression in plants under various stress conditions, but the functional roles of GR-RBPs under stress conditions remain to be verified. Here, we examine the biological roles of a GR-RBP, designated atRZ-1a, in Arabidopsis thaliana under stress conditions. atRZ-1a was expressed ubiquitously in various Arabidopsis organs including stems, roots, leaves, flowers, and siliques. The transcript level of atRZ-1a increased markedly by cold stress, whereas its expression was marginally downregulated by drought stress or abscisic acid treatment. Germination and seedling growth of the loss-of-function mutants were retarded remarkably compared with those of the wild type under cold stress. In contrast, the transgenic Arabidopsis plants that overexpress atRZ-1a displayed earlier germination and better seedling growth than the wild type under cold stress. Moreover, the atRZ-1a-overexpressing transgenic Arabidopsis plants were more freezing tolerant than the wild-type plants. Heterologous expression of atRZ-1a in Escherichia coli demonstrated that the E. coli cells expressing atRZ-1a displayed much higher growth rate than the non-transformed cells after cold shock. These results provide evidence that atRZ-1a affects seed germination and seedling growth under low temperature and plays a role in the enhancement of freezing tolerance in Arabidopsis plants.
DOI:10.1093/pcp/pcm087URLPMID:17602187 [本文引用: 2]
Despite the fact that glycine-rich RNA-binding proteins (GRPs) have been implicated in the responses of plants to changing environmental conditions, the reports demonstrating their biological roles are severely limited. Here, we examined the functional roles of a zinc finger-containing GRP, designated atRZ-1a, in Arabidopsis thaliana under drought or salt stress conditions. Transgenic Arabidopsis plants overexpressing atRZ-1a displayed retarded germination and seedling growth compared with the wild-type plants under salt or dehydration stress conditions. In contrast, the loss-of-function mutants of atRZ-1a germinated earlier and grew faster than the wild-type plants under the same stress conditions. Germination of the transgenic plants and mutant lines was influenced by the addition of ABA or glucose, implying that atRZ-1a affects germination in an ABA-dependent way. H(2)O(2) was accumulated at higher levels in the transgenic plants compared with the wild-type plants under stress conditions. The expression of several germination-responsive genes was modulated by atRZ-1a, and proteome analysis revealed that the expression of different classes of genes, including those involved in reactive oxygen species homeostasis and functions, was affected by atRZ-1a under dehydration or salt stress conditions. Taken together, these results suggest that atRZ-1a has a negative impact on seed germination and seedling growth of Arabidopsis under salt or dehydration stress conditions, and imply that atRZ-1a exerts its function by modulating the expression of several genes under stress conditions.
DOI:10.1105/tpc.15.00949URLPMID:26721863 [本文引用: 2]
Nuclear-localized RNA binding proteins are involved in various aspects of RNA metabolism, which in turn modulates gene expression. However, the functions of nuclear-localized RNA binding proteins in plants are poorly understood. Here, we report the functions of two proteins containing RNA recognition motifs, RZ-1B and RZ-1C, in Arabidopsis thaliana. RZ-1B and RZ-1C were localized to nuclear speckles and interacted with a spectrum of serine/arginine-rich (SR) proteins through their C termini. RZ-1C preferentially bound to purine-rich RNA sequences in vitro through its N-terminal RNA recognition motif. Disrupting the RNA binding activity of RZ-1C with SR proteins through overexpression of the C terminus of RZ-1C conferred defective phenotypes similar to those observed in rz-1b rz-1c double mutants, including delayed seed germination, reduced stature, and serrated leaves. Loss of function of RZ-1B and RZ-1C was accompanied by defective splicing of many genes and global perturbation of gene expression. In addition, we found that RZ-1C directly targeted FLOWERING LOCUS C (FLC), promoting efficient splicing of FLC introns and likely also repressing FLC transcription. Our findings highlight the critical role of RZ-1B/1C in regulating RNA splicing, gene expression, and many key aspects of plant development via interaction with proteins including SR proteins.
DOI:10.1111/j.1365-3040.2009.02101.xURLPMID:20088860 [本文引用: 2]
The rice (Oryza sativa) genome harbours three genes encoding CysCysHisCys (CCHC)-type zinc finger-containing glycine-rich RNA-binding proteins, designated OsRZ proteins, but their importance and physiological functions remain largely unknown. Here, the stress-responsive expression patterns of OsRZs were assessed, and the biological and cellular functions of OsRZs were evaluated under low temperature conditions. The expression levels of the three OsRZs were up-regulated by cold stress, whereas drought or high salt stress did not significantly alter its transcript level. OsRZ2 complemented the cold sensitivity of BX04 Escherichia coli cells under low temperatures, and had DNA-melting activity and transcription anti-termination activity, thereby indicating that OsRZ2 possesses an RNA chaperone activity. By contrast, neither OsRZ1 nor OsRZ3 harboured these activities. Ectopic expression of OsRZ2, but not OsRZ3, in cold-sensitive Arabidopsis grp7 knockout plants rescued the grp7 plants from cold and freezing damage, and OsRZ2 complemented the defect in mRNA export from the nucleus to the cytoplasm in grp7 mutant during cold stress. The present findings support the emerging idea that the regulation of mRNA export is one of the adaptive processes in plants under stress conditions, and RNA chaperone functions as a regulator in mRNA export under cold stress conditions.
DOI:10.1371/journal.pone.0096877URLPMID:24800811 [本文引用: 1]
Although the functional roles of zinc finger-containing glycine-rich RNA-binding proteins (RZs) have been characterized in several plant species, including Arabidopsis thaliana and rice (Oryza sativa), the physiological functions of RZs in wheat (Triticum aestivum) remain largely unknown. Here, the functional roles of the three wheat RZ family members, named TaRZ1, TaRZ2, and TaRZ3, were investigated using transgenic Arabidopsis plants under various abiotic stress conditions. Expression of TaRZs was markedly regulated by salt, dehydration, or cold stress. The TaRZ1 and TaRZ3 proteins were localized to the nucleus, whereas the TaRZ2 protein was localized to the nucleus, endoplasmic reticulum, and cytoplasm. Germination of all three TaRZ-expressing transgenic Arabidopsis seeds was retarded compared with that of wild-type seeds under salt stress conditions, whereas germination of TaRZ2- or TaRZ3-expressing transgenic Arabidopsis seeds was retarded under dehydration stress conditions. Seedling growth of TaRZ1-expressing transgenic plants was severely inhibited under cold or salt stress conditions, and seedling growth of TaRZ2-expressing plants was inhibited under salt stress conditions. By contrast, expression of TaRZ3 did not affect seedling growth of transgenic plants under any of the stress conditions. In addition, expression of TaRZ2 conferred freeze tolerance in Arabidopsis. Taken together, these results suggest that different TaRZ family members play various roles in seed germination, seedling growth, and freeze tolerance in plants under abiotic stress.
DOI:10.1016/S0968-0004(98)01255-9URLPMID:9757828 [本文引用: 1]
Abstract Members of a family of cold-shock proteins (CSPs) are found throughout the eubacterial domain and appear to function as RNA-chaperones. They have been implicated in various cellular processes, including adaptation to low temperatures, cellular growth, nutrient stress and stationary phase. The discovery of a domain--the cold-shock domain--that shows strikingly high homology and similar RNA-binding properties to CSPs in a growing number of eukaryotic nucleic-acid-binding proteins suggests that these proteins have an ancient origin.
DOI:10.1016/j.bbrc.2014.11.092URLPMID:25475723 [本文引用: 2]
The cold shock domain (CSD) is the most conserved nucleic acid binding domain and is distributed from bacteria to animals and plants. CSD proteins are RNA chaperones that destabilize RNA secondary structures to regulate stress tolerance and development. AtCSP2 is one of the four CSD proteins in Arabidopsis and is up-regulated in response to cold. Since AtCSP2 negatively regulates freezing tolerance, it was proposed to be a modulator of freezing tolerance during cold acclimation. Here, we examined the function of AtCSP2 in seed germination. We found that AtCSP2-overexpressing lines demonstrated retarded germination as compared with the wild type, with or without stress treatments. The ABA levels in AtCSP2-overexpressing seeds were higher than those in the wild type. In addition, overexpression of AtCSP2 reduced the expression of an ABA catabolic gene (CYP707A2) and gibberellin biosynthesis genes (GA20ox and GA3ox). These results suggest that AtCSP2 negatively regulates seed germination by controlling ABA and GA levels.
DOI:10.1093/pcp/pcp037URLPMID:19258348 [本文引用: 1]
Unlike the well-known functions of cold shock proteins in prokaryotes during cold adaptation, the biological functions of cold shock domain proteins (CSDPs) in plants remain largely unknown. Here, we examined the functional roles of two structurally different CSDPs, CSDP1 harboring a long C-terminal glycine-rich region interspersed with seven CCHC-type zinc fingers and CSDP2 containing a far shorter glycine-rich region interspersed with two CCHC-type zinc fingers, in Arabidopsis thaliana under stress conditions. CSDP1 overexpression delayed the seed germination of Arabidopsis under dehydration or salt stress conditions, whereas CSDP2 overexpression accelerated the seed germination of Arabidopsis under salt stress conditions. CSDP1 and CSDP2 rescued the cold-sensitive glycine-rich RNA-binding protein 7 mutant plants from freezing damage to a different degree, and this rescuing capability was correlated with their ability to complement the cold-sensitive Escherichia coli BX04 mutant at low temperatures. The nucleic acid-binding properties of CSDPs varied depending on the N-terminal cold shock domain and the C-terminal glycine-rich zinc finger region. Collectively, these results showed that CSDP1 and CSDP2 perform different functions in seed germination and growth of Arabidopsis under stress conditions, and that the glycine-rich region interspersed with CCHC-type zinc fingers is particularly important for its nucleic acid-binding activities and function.
DOI:10.1111/nph.12118URLPMID:23323758 [本文引用: 1]
Bacterial cold shock proteins (CSPs) act as RNA chaperones that destabilize mRNA secondary structures at low temperatures. Bacterial CSPs are composed solely of a nucleic acid-binding domain termed the cold shock domain (CSD). Plant CSD proteins contain an auxiliary domain in addition to the CSD but also show RNA chaperone activity. However, their biological functions are poorly understood.We examined Arabidopsis COLD SHOCK DOMAIN PROTEIN 2 (AtCSP2) using overexpressing and mutant lines.A double mutant, with reduced AtCSP2 and no AtCSP4, showed higher freezing tolerance than the wild-type when cold-acclimated. The increase in freezing tolerance was associated with up-regulation of CBF transcription factors and their downstream genes. By contrast, overexpression of AtCSP2 resulted in decreased freezing tolerance when cold-acclimated. In addition, late flowering and shorter siliques were observed in the overexpressing lines.AtCSP2 negatively regulates freezing tolerance and is partially redundant with its closest paralog, AtCSP4.
DOI:10.1016/j.fob.2013.10.003URLPMID:3829988 [本文引用: 1]
61Arabidopsis thaliana COLD SHOCK DOMAIN PROTEIN 3 (AtCSP3) is induced during salt and drought stresses.61A knockout mutant of AtCSP3 showed lower survival rates after salt and drought stresses.61AtCSP3-overexpressing plants displayed higher survival rates after salt and drought stresses.61AtCSP3 is involved in the regulation of salt and drought stress tolerance in Arabidopsis.
[本文引用: 1]
DOI:10.1016/j.plaphy.2014.01.004URLPMID:24534004 [本文引用: 2]
61Accumulation of wheat cold shock domain protein family was serologically analyzed.61Four WCSP members were identified and classified into three classes.61Class I and II WCSPs may be involved in cold adaptation in meristematic tissues.61Class III WCSP may be involved in starch filling in seeds.
DOI:10.1111/j.1365-3040.2008.01811.xURLPMID:18397370 [本文引用: 2]
Two novel rice cold shock domain (CSD) proteins were cloned and characterized under different stress treatments and during various stages of development. OsCSP1 and OsCSP2 ( Oryza sativa CSD protein) encode putative proteins consisting of an N-terminal CSD and glycine-rich regions that are interspersed by 4 and 2 C X 2 C X 4 H X 4 C (CCHC) retroviral-like zinc fingers, respectively. In vivo functional analysis confirmed that OsCSPs can complement a cold-sensitive bacterial strain which lacks four endogenous cold shock proteins. In vitro ssDNA binding assays determined that recombinant OsCSPs are capable of functioning as nucleic acid-binding proteins. Both OsCSP transcripts are transiently up-regulated in response to low-temperature stress and rapidly return to a basal level of gene expression. Protein blot analysis determined that OsCSPs are maintained at a constant level subsequent to a cold treatment lasting over a period of several days. Both the transcript and protein data are in sharp contrast to those previously obtained for winter wheat WCSP1. A time-coursed study through various stages of rice development confirmed that both OsCSP proteins and transcripts are highly accumulated in reproductive tissues and tissues which exhibit meristematic activity.
DOI:10.1016/j.biochi.2015.04.004URLPMID:25882680 [本文引用: 1]
61Pentatricopeptide repeat (PPR) proteins regulate gene expression at the RNA level.61PPR proteins bind RNA in a sequence-specific, modular fashion.61Designer PPR proteins can be constructed to influence gene expression in02vivo.61Loss of PPR proteins in eukaryotes can have deleterious consequences.61Horizontal gene transfer has played a key role in the distribution of PPR proteins.
DOI:10.1038/s41598-018-21269-1URL [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins, which are characterized by tandem 30 40 amino acid sequence motifs, constitute of a large gene family in plants. Some PPR proteins have been identified to play important roles in organellar RNA metabolism and organ development inArabidopsisand rice. However, functions ofPPRgenes in woody species remain largely unknown. Here, we identified and characterized a total of 626PPRgenes containing PPR motifs in thePopulus trichocarpagenome. A comprehensive genome-wide analysis of the poplarPPRgene family was performed, including chromosomal location, phylogenetic relationships and gene duplication. Genome-wide transcriptomic analysis showed that 154 of thePtrPPRgenes were induced by biotic and abiotic treatments, includingMarssonina brunnea, salicylic acid (SA), methyl jasmonate (MeJA), mechanical wounding, cold and salinity stress. Quantitative RT-PCR analysis further investigated the expression profiles of 11PtrPPRgenes under different stresses. Our results contribute to a comprehensive understanding the roles of PPR proteins and provided an insight for improving the stress tolerance in poplar.
DOI:10.1016/j.jgg.2018.01.006URLPMID:29580769 [本文引用: 1]
In plants, RNA editing is a post-transcriptional process that changes specific cytidine to uridine in both mitochondria and plastids. Most pentatricopeptide repeat (PPR) proteins are involved in organelle RNA editing by recognizing specific RNA sequences. We here report the functional characterization of a PPR protein from the DYW subclass, Baili Xi (BLX), which contains five PPR motifs and a DYW domain. BLX is essential for early seed development, as plants lacking the BLX gene was embryo lethal and the endosperm failed to initiate cellularization. BLX was highly expressed in the embryo and endosperm, and the BLX protein was specifically localized in mitochondria, which is essential for BLX function. We found that BLX was required for the efficient editing of 36 editing sites in mitochondria. Moreover, BLX was involved in the splicing regulation of the fourth intron of nad1 and the first intron of nad2 . The loss of BLX function impaired the mitochondrial function and increased the reactive oxygen species (ROS) level. Genetic complementation with truncated variants of BLX revealed that, in addition to the DYW domain, only the fifth PPR motif was essential for BLX function. The upstream sequences of the BLX-targeted editing sites are not conserved, suggesting that BLX serves as a novel and major mitochondrial editing factor (MEF) via a new non-RNA-interacting manner. This finding provides new insights into how a DYW-type PPR protein with fewer PPR motifs regulates RNA editing in plants.
DOI:10.1093/jxb/erw331URL [本文引用: 1]
The mitochondrial pentatricopeptide repeat protein AtGRS1 edits RNA at four sites and is critical for development. The abscisic acid response geneABI5participates in the short root phenotype ofgrs1. Most pentatricopeptide repeat (PPR) proteins are involved in organelle post-transcriptional processes, including RNA editing. The PPR proteins include the PLS subfamily, containing characteristic triplets of P, L, and S motifs; however, their editing mechanisms and roles in developmental processes are not fully understood. In this study, we isolated theArabidopsis thaliana Growing slowly 1(AtGRS1) gene and showed that it functions in RNA editing and plant development. Arabidopsis null mutants ofgrs1exhibit slow growth and sterility. Further analysis showed that cell division activity was reduced dramatically in the roots ofgrs1plants. We determined that GRS1 is a nuclear-encoded mitochondria-localized PPR protein, and is a member of the PLS subfamily. GRS1 is responsible for the RNA editing at four specific sites of four mitochondrial mRNAs:nad1-265,nad4L-55,nad6-103, andrps4-377. The first three of these mRNAs encode for the subunits of complex I of the electron transport chain in mitochondria. Thus, the activity of complex I is strongly reduced ingrs1. Changes inRPS4editing ingrs1plants affect mitochondrial ribosome biogenesis. Expression of the alternative respiratory pathway and the abscisic acid response geneABI5were up-regulated ingrs1mutant plants. Genetic analysis revealed thatABI5is involved in the short root phenotype ofgrs1. Taken together, our results indicate thatAtGRS1regulates plant development by controlling RNA editing in Arabidopsis.
DOI:10.1007/s11103-016-0566-4URLPMID:27942959 [本文引用: 1]
Abstract KEY MESSAGE: SLO4 is a mitochondrial PPR protein that is involved in editing nad4, possibly required for the efficient splicing of nad2 intron1. Pentatricopeptide repeat (PPR) proteins constitute a large protein family in flowering plants and are thought to be mostly involved in organellar RNA metabolism. The subgroup of PLS-type PPR proteins were found to be the main specificity factors of cytidine to uridine RNA editing. Identifying the targets of PLS-type PPR proteins can help in elucidating the molecular function of proteins encoded in the organellar genomes. In this study, plants lacking the SLOW GROWTH 4 PPR protein were characterized. Slo4 mutants were characterized as having restricted root growth, being late flowering and displaying an overall delayed growth phenotype. Protein levels and activity of mitochondrial complex I were decreased and putative complex I assembly intermediates accumulated in the mutant plants. An editing defect, leading to an amino acid change, in the mitochondrial nad4 transcript, encoding for a complex I subunit, was identified. Furthermore, the splicing efficiency of the first intron of nad2, encoding for another complex I subunit, was also decreased. The change in splicing efficiency could however not be linked to any editing defects in the nad2 transcript.
DOI:10.1111/tpj.12143URLPMID:23398165 [本文引用: 1]
The 5 ends of many mitochondrial transcripts are generated post-transcriptionally. Recently, we identified three RNA PROCESSING FACTORs required for 5 end maturation of different mitochondrial mRNAs in Arabidopsis thaliana. All of these factors are pentatricopeptide repeat proteins (PPRPs), highly similar to RESTORERs OF FERTILTY (RF), that rescue male fertility in cytoplasmic male-sterile lines from different species. Therefore, we suggested a general role of these RF-like PPRPs in mitochondrial 5 processing. We now identified RNA PROCESSING FACTOR 5, a PPRP not classified as an RF-like protein, required for the efficient 5 maturation of the nad6 and atp9 mRNAs as well as 26S rRNA. The precursor molecules of these RNAs share conserved sequence elements, approximately ranging from positions 50 to +9 relative to mature 5 mRNA termini, suggesting these sequences to be at least part of the cis elements required for processing. The knockout of RPF5 has only a moderate influence on 5 processing of atp9 mRNA, whereas the generation of the mature nad6 mRNA and 26S rRNA is almost completely abolished in the mutant. The latter leads to a 50% decrease of total 26S rRNA species, resulting in an imbalance between the large rRNA and 18S rRNA. Despite these severe changes in RNA levels and in the proportion between the 26S and 18S rRNAs, mitochondrial protein levels appear to be unaltered in the mutant, whereas seed germination capacity is markedly reduced.
[本文引用: 1]
DOI:10.1093/jxb/eru293URLPMID:4157714 [本文引用: 1]
The authors identify a pentatricopeptide repeat (PPR) protein SOAR1 as a crucial player of ABA signalling, which localizes to both the cytosol and nucleus probably to regulate nuclear gene expression. A dominant suppressor of the ABAR overexpressor, soar1-1D, from CHLH/ABAR [coding for Mg-chelatase H subunit/putative abscisic acid (ABA) receptor (ABAR)] overexpression lines was screened to explore the mechanism of the ABAR-mediated ABA signalling. The SOAR1 gene encodes a pentatricopeptide repeat (PPR) protein which localizes to both the cytosol and nucleus. Down-regulation of SOAR1 strongly enhances, but up-regulation of SOAR1 almost completely impairs, ABA responses, revealing that SOAR1 is a critical, negative, regulator of ABA signalling. Further genetic evidence supports that SOAR1 functions downstream of ABAR and probably upstream of an ABA-responsive transcription factor ABI5. Changes in the SOAR1 expression alter expression of a subset of ABA-responsive genes including ABI5. These findings provide important information to elucidate further the functional mechanism of PPR proteins and the complicated ABA signalling network.
DOI:10.1111/j.1365-313X.2011.04883.xURLPMID:22248025 [本文引用: 1]
Land plants contain a large family of genes that encode for pentatricopeptide (PPR) proteins. To date, few of these PPR proteins have been functionally characterized. In this study, we have analyzed an Arabidopsis mutant, slg1, which exhibits slow growth and delayed development. In addition, slg1 shows an enhanced response to ABA and increased tolerance to drought stress. The SLG1 gene encodes a PPR protein that is localized in mitochondria. In the slg1 mutant, RNA editing in a single site of the mitochondrial transcript nad3 is abolished. nad3 is a subunit of complex I of the electron transport chain in mitochondria. As a consequence, the NADH dehydrogenase activity of complex I in slg1 is strongly impaired and production of ATP is reduced. When responding to ABA treatment, slg1 accumulates more H2O2 in its guard cells than the wild type. The slg1 mutant also has an increased expression of genes involved in the alternative respiratory pathway, which may compensate for the disrupted function of complex I and help scavenge the excess accumulation of H2O2. Our functional characterization of the slg1 mutant revealed a putative link between mitochondrial RNA editing and plant responses to abiotic stress.
DOI:10.1104/pp.111.177501URLPMID:21653783 [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins (PPRPs) are encoded by a large gene family in Arabidopsis (Arabidopsis thaliana), and their functions are largely unknown. The few studied PPRPs are implicated in different developmental processes through their function in RNA metabolism and posttranscriptional regulation in plant organelles. Here, we studied the functions of Arabidopsis PENTATRICOPEPTIDE REPEAT PROTEIN FOR GERMINATION ON NaCl (PGN) in plant defense and abiotic stress responses. Inactivation of PGN results in susceptibility to necrotrophic fungal pathogens as well as hyper sensitivity to abscisic acid (ABA), glucose, and salinity. Interestingly, ectopic expression of PGN results in the same phenotypes as the pgn null allele, indicating that a tight regulation of the PGN transcript is required for normal function. Loss of PGN function dramatically enhanced reactive oxygen species accumulation in seedlings in response to salt stress. Inhibition of ABA synthesis and signaling partially alleviates the glucose sensitivity of pgn, suggesting that the mutant accumulates high endogenous ABA. Accordingly, induction of NCED3, encoding the rate-limiting enzyme in stress-induced ABA biosynthesis, is significantly higher in pgn, and the mutant has higher basal ABA levels, which may underlie its phenotypes. The pgn mutant has altered expression of other ABA-related genes as well as mitochondria-associated transcripts, most notably elevated levels of ABI4 and ALTERNATIVE OXIDASEla, which are known for their roles in retrograde signaling induced by changes in or inhibition of mitochondrial function. These data, coupled with its mitochondrial localization, suggest that PGN functions in regulation of reactive oxygen species homeostasis in mitochondria during abiotic and biotic stress responses, likely through involvement in retrograde signaling.
DOI:10.1104/pp.113.227199URLPMID:24144791 [本文引用: 1]
Chloroplasts are the site of photosynthesis and the biosynthesis of essential metabolites, including amino acids, fatty acids, and secondary metabolites. It is known that many seedling-lethal mutants are impaired in chloroplast function or development, indicating the development of functional chloroplast is essential for plant growth and development. Here, we isolated a novel transfer DNA insertion mutant, dubbed sel1 (for seedling lethal1), that exhibited a pigment-defective and seedling-lethal phenotype with a disrupted pentatricopeptide repeat (PPR) gene. Sequence analysis revealed that SEL1 is a member of the PLS subgroup, which is lacking known E/E+ or DYW domains at the C terminus, in the PLS subfamily of the PPR protein family containing a putative N-terminal transit peptide and 14 putative PPR or PPR-like motifs. Confocal microscopic analysis showed that the SEL1-green fluorescent protein fusion protein is localized in chloroplasts. Transmission electron microscopic analysis revealed that the sel1 mutant is impaired in the etioplast, as well as in chloroplast development. In sel1 mutants, plastid-encoded proteins involved in photosynthesis were rarely detected due to the lack of the corresponding transcripts. Furthermore, transcript profiles of plastid genes revealed that, in sel1 mutants, the transcript levels of plastid-encoded RNA polymerase-dependent genes were greatly reduced, but those of nuclear-encoded RNA polymerase-dependent genes were increased or not changed. Additionally, the RNA editing of two editing sites of the acetyl-CoA carboxylase beta subunit gene transcripts in the sel1 mutant was compromised, though it is not directly connected with the sel1 mutant phenotype. Our results demonstrate that SEL1 is involved in the regulation of plastid gene expression required for normal chloroplast development.
DOI:10.1111/j.1365-313X.2011.04569.xURLPMID:21435048 [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins are mainly involved in regulating post-transcriptional processes in mitochondria and plastids, including chloroplasts. Mutations in the Arabidopsis PPR2 gene have previously been found to cause defects in seed development and reduced transmission through male and female gametophytes. However, the exact function of AtPPR2 has not been defined. We found that a loss-of-function mutation of AtPPR2 leads to arrest of the first mitotic division during both male and female gametogenesis. In addition, the Atppr2 mutation causes delayed embryogenesis, leading to embryonic lethality. Mutation in emb2750, which appears to be a weak mutant allele of the AtPPR2 locus, also results in defective seeds. However, a majority of emb2750 seeds were able to germinate, but their cotyledons were albino and often deformed, and growth of the emb2750 seedlings were arrested after germination. AtPPR2 is mainly expressed in plant parts that undergo cell division, and AtPPR2 protein was localized to chloroplasts. RNA immunoprecipitation and protein gel mobility shift assays showed that AtPPR2 binds to plastid 23S rRNA. Our study adds to a growing body of evidence that plastids and/or chloroplasts play a key role in cell division. AtPPR2 may modulate the translational process to fine-tune plastid function, thereby regulating cell division.
DOI:10.1007/s10265-012-0527-1URLPMID:23076438 [本文引用: 1]
AbstractRNA processing, RNA editing, RNA splicing and translational activation of RNAs are essential post-transcriptional steps in chloroplast gene expression. Typically, the factors mediating those processes are nuclear encoded and post-translationally imported into the chloroplasts. In land plants, members of the large pentatricopeptide repeat (PPR) protein family are required for individual steps in chloroplast RNA processing. Interestingly, a subgroup of PPR proteins carries a C-terminal small MutS related (SMR) domain. Here we analyzed the consequences of mutations in the gene, which encodes a PPR-SMR protein, in mutations lead to a specific reduction in chloroplast ATP synthase levels. Furthermore, we found aberrant transcript patterns for ATP synthase coding mRNAs in mutants. Finally, a reduced ribosome association of / and mRNAs in mutants suggests the involvement of the PPR-SMR protein SVR7 in translational activation of these mRNAs. We describe that the function of SVR7 in translation has expanded relative to its maize ortholog ATP4. The results provide evidence for a relaxed functional conservation of this PPR-SMR protein in eudicotyledonous and monocotyledonous plants, thus adding to the knowledge about the function and evolution of PPR-SMR proteins.
DOI:10.1038/s41598-017-10727-xURLPMID:5597598 [本文引用: 1]
Abstract Several mitochondrial-targeted pentatricopeptide repeat (PPR) proteins involved in pollen development have been reported to be fertility restorer (Rf) proteins. However, the roles of plastid-localized PPR proteins in plant male reproduction are poorly defined. Here, we described a plastid-localized PPR-SMR protein, OsPPR676, which is required for plant growth and pollen development in rice. In this study, OsPPR676 was confirmed to be an interacted protein with Osj10gBTF3, -subunit of nascent polypeptide-associated complex (-NAC), by bimolecular fluorescence complementation assays, indicating that both proteins are probably involved in the same regulatory pathway of pollen development. Compared with other chloroplast-rich tissues, OsPPR676 was only weakly expressed in anther, but in the Mei and YM stages of pollen development, its expression was relatively strong in the tapetum. Disruption of OsPPR676 resulted in growth retardation of plants and partial sterility of pollens. Phenotypic analysis of different osppr676 mutant lines implied that the SMR domain was not essential for the function of OsPPR676. We further demonstrated that OsPPR676 is essential for production of plastid atpB subunit, and then plays crucial roles in biosynthesis of fatty acids, carbohydrates, and other organic matters via affecting activity of ATP synthase.
DOI:10.1007/s11103-017-0654-0URLPMID:28856519 [本文引用: 1]
OsPPR6, a pentatricopeptide repeat protein involved in editing and splicing chloroplast RNA, is required for chloroplast biogenesis in rice. The chloroplast has its own genetic material and genetic sy
DOI:10.1093/jxb/ery108URLPMID:29562289 [本文引用: 1]
Abstract In flowering plants, various RNA editing events occur in the mitochondria and chloroplasts as part of post-transcriptional processes. Although several PPRs and MORFs have been identified as RNA editing factors, the underlying mechanism of PPRs and the cooperation among these proteins are still obscure. Here, we identified a rice dual-localized PPR protein, OsPGL1. The loss of function of OsPGL1 resulted in defects in both chloroplast RNA editing of ndhD-878 and mitochondrial RNA editing of ccmFc-543, both of which could be restored via complementary validation. Despite synonymous editing of ccmFc-543, the loss of editing of ndhD-878 caused a failed conversion of serine to leucine, leading to chloroplast dysfunction and defects in the photosynthetic complex; the results of additional experiments demonstrated that OsPGL1 directly binds to both transcripts. Interactions between three OsMORFs (OsMORF2/8/9) and OsPGL1 both in vitro and in vivo were confirmed, implying that OsPGL1 functions in RNA editing via an editosome. These findings also suggested that OsMORFs assist with and contribute to a flexible PPR-RNA recognition model during RNA editing. These results indicate that, in cooperation with PPRs, OsPGL1 is required for RNA editing. In addition, these results provide new insight into the relationship between RNA editing and plant development.
DOI:10.1105/tpc.111.093211URLPMID:22247252 [本文引用: 1]
The cytoplasmic male sterility (CMS) phenotype in plants can be reversed by the action of nuclear-encoded fertility restorer (Rf) genes. The molecular mechanism involved in Rf gene-mediated processing of CMS-associated transcripts is unclear, as are the identities of other proteins that may be involved in the CMS-Rf interaction. In this study, we cloned the restorer gene Rf5 for Hong-Lian CMS in rice and studied its fertility restoration mechanism with respect to the processing of the CMS-associated transcript atp6-orfH79. RF5, a pentatricopeptide repeat (PPR) protein, was unable to bind to this CMS-associated transcript; however, a partner protein of RF5 (GRP162, a Gly-rich protein encoding 162 amino acids) was identified to bind to atp6-orfH79. GRP162 was found to physically interact with RF5 and to bind to atp6-orfH79 via an RNA recognition motif. Furthermore, we found that RF5 and GRP162 are both components of a restoration of fertility complex (RFC) that is 400 to 500 kD in size and can cleave CMS-associated transcripts in vitro. Evidence that a PPR protein interacts directly with a Gly-rich protein to form a subunit of the RFC provides a new perspective on the molecular mechanisms underlying fertility restoration.
DOI:10.1093/mp/ssu054URLPMID:24821718 [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins play a central role in modulating organellar gene expression by participating in various aspects of organellar RNA metabolism. In this study, we identify a novel rice PPR protein WSL, which is involved in the splicing of chloroplast rpl2 and thereby affects chloroplast development and abiotic stress response in rice.
DOI:10.3389/fpls.2017.01116URLPMID:5483476 [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins comprise a large family in higher plants and perform diverse functions in organellar RNA metabolism. Despite the rice genome encodes 477 PRR proteins, the regulatory effects of PRR proteins on chloroplast development remains unknown. In this study, we report the functional characterization of the ricewhite stripe leaf4(wsl4) mutant. Thewsl4mutant develops white-striped leaves during early leaf development, characterized by decreased chlorophyll content and malformed chloroplasts. Positional cloning of theWSL4gene, together with complementation and RNA-interference tests, reveal that it encodes a novel P-family PPR protein with 12 PPR motifs, and is localized to chloroplast nucleoids. Quantitative RT-PCR analyses demonstrate thatWSL4is a low temperature response gene abundantly expressed in young leaves. Further expression analyses show that many nuclear- and plastid-encoded genes in thewsl4mutant are significantly affected at the RNA and protein levels. Notably, thewsl4mutant causes defects in the splicing ofatpF, ndhA, rpl2, andrps12. Our findings identify WSL4 as a novel P-family PPR protein essential for chloroplast RNA group II intron splicing during early leaf development in rice.
DOI:10.1111/jipb.12138URLPMID:24289830 [本文引用: 1]
Pentatricopeptide repeat (PPR) proteins, characterized by tandem arrays of a 35 amino acid motif, have been suggested to play central and broad roles in modulating the expression of organelle genes in plants. However, the molecular mechanisms of most rice PPR genes remains unclear. In this paper, we isolated and characterized a temperature-conditional virescent mutant, OsV4, in rice (Oryza sativa cultivar Jiahua1 (WT, japonica rice variety)). The mutant displays albino phenotype and abnormal chloroplasts at the three leaf stage, which gradually turns green after the four leaf stage at a low temperature (2009000900°C). But the mutant always develops green leaves and well-developed chloroplasts at a high temperature (3209000900°C). Genetic and molecular analyses uncovered that OsV4 encodes a novel chloroplast-targeted PPR protein including four PPR motifs. Further investigations show that the mutant phenotype is associated with changes in chlorophyll content and chloroplast development. The OsV4 transcripts only accumulate to high levels in young leaves, indicating that its expression is tissue-specific. In addition, transcript levels of some ribosomal components and plastid-encoded polymerase-dependent genes are dramatically reduced in the albino mutants grown at 2009000900°C. These findings suggest that OsV4 plays an important role during early chloroplast development under cold stress in rice.
DOI:10.1111/tpj.13551URLPMID:28346745 [本文引用: 1]
In higher plants, many mitochondrial genes contain group II‐type introns that are removed from RNAs by splicing to produce mature transcripts that are then translated into functional proteins. However, the factors involved in the splicing of mitochondrial introns and their biological functions are not well understood in maize. Here, we isolated an () mutant and identified the underlying gene by map‐based cloning. encodes a P‐type mitochondria‐targeted pentatricopeptide repeat (PPR) protein with 10 PPR motifs. Loss of function results in splicing defect of the first intron of , a gene encoding subunit 2 of NADH dehydrogenase (also called complex I). The mutant has undetectable activity of complex I and has arrested development of embryo and endosperm, and thus defective seeds with empty pericarp. Additionally, the basal endosperm transfer layer cells were severely affected, indicating the deficiency of cell wall ingrowths in the kernels. Moreover, the alternative respiratory pathway involving alternative oxidase was significantly induced in the mutant. These results suggest that EMP10 is specifically required for the ‐splicing of mitochondrial intron 1, embryogenesis and endosperm development in maize.
DOI:10.1093/jxb/erx212URLPMID:28981788 [本文引用: 1]
A new pentatricopeptide repeat (PPR) gene in maize differs from previously reported genes, as it is required for splicing of all four introns ofnad1. Group II introns are common in the mitochondrial genome of higher plant species. The splicing of these introns is a complex process involving the synergistic action of multiple factors. However, few of these factors have been characterized in maize. In this study, we found that theEmpty pericarp11(Emp11) gene, which encodes a P-type pentatricopeptide repeat (PPR) protein, is required for the development of maize seeds. The loss ofEmp11function seriously impairs embryo and endosperm development, resulting in empty pericarp seeds in maize, and alteration inEmp11expression leads to quantitative variation in kernel size and weight. We found that theemp11mutants showed a failure innad1intron splicing, exhibited a severe reduction in complex I assembly and activity, mitochondrial structure disturbances, and an increase in alternative oxidaseAOX2andAOX3levels. Interestingly, theemp11phenotype was very severe in the W22 inbred line but could be partially recovered in B73 BC2F2segregating ears. These results suggest that EMP11 serves as a factor for the splicing of mitochondrialnad1introns and is required for mitochondrial function to ensure proper seed development in maize.
DOI:10.1534/genetics.116.199331URLPMID:28213476 [本文引用: 1]
Respiration, the core of mitochondrial metabolism, depends on the function of five respiratory complexes. Many respiratory chain-related proteins are encoded by the mitochondrial genome and their RNAs undergo post-transcriptional modifications by nuclear genome-expressed factors, including pentatricopeptide repeat (PPR) proteins. Maize defective kernel 10 (dek10) is a classic mutant with small kernels and delayed development. Through positional cloning, we found that Dek10 encodes an E-subgroup PPR protein localized in mitochondria. Sequencing analysis indicated that Dek10 is responsible for the C-to-U editing at nad3-61, nad3-62, and cox2-550 sites, which are specific editing sites in monocots. The defects of these editing sites result in significant reduction of Nad3 and the loss of Cox2. Interestingly, the assembly of complex I was not reduced, but its NADH dehydrogenase activity was greatly decreased. The assembly of complex IV was significantly reduced. Transcriptome and transmission electron microscopy (TEM) analysis revealed that proper editing of nad3 and cox2 is critical for mitochondrial functions, biogenesis, and morphology. These results indicate that the E-subgroup PPR protein Dek10 is responsible for multiple editing sites in nad3 and cox2, which are essential for mitochondrial functions and plant development in maize.
DOI:10.1016/j.molp.2016.08.008URLPMID:27596292 [本文引用: 1]
DOI:10.1016/j.molp.2017.09.009URL [本文引用: 1]
DOI:10.1105/tpc.111.091074URL [本文引用: 1]
RNA editing plays an important role in organelle gene expression in various organisms, including flowering plants, changing the nucleotide information at precise sites. Here, we present evidence that the maize (Zea mays) nuclear gene Pentatricopeptide repeat 2263 (PPR2263) encoding a DYW domain-containing PPR protein is required for RNA editing in the mitochondrial NADH dehydrogenase5 (nad5) and cytochrome b (cob) transcripts at the nad5-1550 and cob-908 sites, respectively. Its putative ortholog, MITOCHONDRIAL EDITING FACTOR29, fulfills the same role in Arabidopsis thaliana. Both the maize and the Arabidopsis proteins show preferential localization to mitochondria but are also detected in chloroplasts. In maize, the corresponding ppr2263 mutation causes growth defects in kernels and seedlings. Embryo and endosperm growth are reduced, leading to the production of small but viable kernels. Mutant plants have narrower and shorter leaves, exhibit a strong delay in flowering time, and generally do not reach sexual maturity. Whereas mutant chloroplasts do not have major defects, mutant mitochondris lack complex III and are characterized by a compromised ultrastructure, increased transcript levels, and the induction of alternative oxidase. The results suggest that mitochondrial RNA editing at the cob-908 site is necessary for mitochondrion biogenesis, cell division, and plant growth in maize.
DOI:10.1007/s11103-009-9584-9URLPMID:19960362 [本文引用: 1]
In Arabidopsis thaliana the putative mitochondrial RNA helicases PMH1 and PMH2 are members of the large DEAD-box protein family. Our previous characterization of these proteins revealed that PMH1 and/or PMH2 are part of high molecular weight complexes. Now T-DNA insertion lines were established and characterized for each of these genes. Immunodetection analysis of cell suspension cultures established from pmh1-1 and pmh2-1 mutants revealed that indeed both DEAD-box proteins are detectable in large protein complexes with PMH2 being much more abundant than PMH1. In plants the knockout of PMH2 leads to reduced group II intron splicing efficiency. In addition the steady-state levels of several mature mitochondrial mRNAs are decreased while transcription is not influenced. This molecular phenotype suggests that PMH2 acts at the posttranscriptional level with a potential function as RNA chaperone required for formation or maintenance of complex RNA secondary structures of introns rather than a direct role in splicing. In contrast, the investigation of a pmh1-1 knockout line did not reveal any influence of this protein on processing and abundance of mitochondrial transcripts.
DOI:10.1104/pp.17.01545URLPMID:29138350 [本文引用: 1]
Abstract DEAD-box RNA helicases (DBRHs) modulate RNA secondary structure, allowing RNA molecules to adopt the conformations required for interaction with their target proteins. RH50 is a chloroplast-located DBRH that co-localizes and is co-expressed with GUN1, a central factor in chloroplast-to-nucleus signaling. When combined with mutations that impair plastid gene expression (prors1-1, prpl11-1, prps1-1, prps21-1, prps17-1 and prpl24-1), rh50 and gun1 mutations evoke similar patterns of epistatic effects. These observations, together with the synergistic growth phenotype of the double mutant rh50-1 gun1-102, suggest that RH50 and GUN1 are functionally related and that this function is associated with plastid gene expression, in particular ribosome functioning. However, rh50-1 itself is not a gun mutant, although - like gun1-102 - the rh50-1 mutation suppresses the down-regulation of nuclear genes for photosynthesis induced by the prors1-1 mutation. The RH50 protein co-migrates with ribosomal particles, and is required for efficient translation of plastid proteins. RH50 binds to transcripts of the 23S-4.5S intergenic region and, in its absence, levels of the corresponding rRNA processing intermediate are strongly increased, implying that RH50 is required for the maturation of the 23S and 4.5S rRNAs. This inference is supported by the finding that loss of RH50 renders chloroplast protein synthesis sensitive to erythromycin and exposure to cold. Based on these results, we conclude that RH50 is a plastid rRNA maturation factor. {copyright, serif} 2017 American Society of Plant Biologists. All rights reserved.
DOI:10.1104/pp.112.197525URLPMID:22576849 [本文引用: 1]
Chloroplasts in angiosperms contain at least seven nucleus-encoded members of the DEAD box RNA helicase family. Phylogenetic analysis shows that five of these plastid members (RH22, -39, -47, -50, and -58) form a single clade and that RH3 forms a clade with two mitochondrial RH proteins (PMH1 and -2) functioning in intron splicing. The function of chloroplast RH3 in maize (Zea mays; ZmRH3) and Arabidopsis (Arabidopsis thaliana; AtRH3) was determined. ZmRH3 and AtRH3 are both under strong developmental control, and ZmRH3 abundance sharply peaked in the sink-source transition zone of developing maize leaves, coincident with the plastid biogenesis machinery. ZmRH3 coimmunoprecipitated with a specific set of plastid RNAs, including several group II introns, as well as pre23S and 23S ribosomal RNA (rRNA), but not 16S rRNA. Furthermore, ZmRH3 associated with 50S preribosome particles as well as nucleoids. AtRH3 null mutants are embryo lethal, whereas a weak allele (rh3-4) results in pale-green seedlings with defects in splicing of several group II introns and rRNA maturation as well as reduced levels of assembled ribosomes. These results provide strong evidence that RH3 functions in the splicing of group II introns and possibly also contributes to the assembly of the 50S ribosomal particle. Previously, we observed 5-to 10-fold upregulation of AtRH3 in plastid Caseinolytic protease mutants. The results shown here indicate that AtRH3 up-regulation was not a direct consequence of reduced proteolysis but constituted a compensatory response at both RH3 transcript and protein levels to impaired chloroplast biogenesis; this response demonstrates that cross talk between the chloroplast and the nucleus is used to regulate RH3 levels.
DOI:10.1105/tpc.112.108340URLPMID:23371945 [本文引用: 2]
Cold stress resulting from chilling and freezing temperatures substantially reduces crop production worldwide. To identify genes critical for cold tolerance in plants, we screened Arabidopsis thaliana mutants for deregulated expression of a firefly luciferase reporter gene under the control of the C-REPEAT BINDING FACTOR2 (CBF2) promoter (CBF2:LUC). A regulator of CBF gene expression1 (rcf1-1) mutant that is hypersensitive to cold stress was chosen for in-depth characterization. RCF1 encodes a cold-inducible DEAD (Asp-Glu-Ala-Asp) box RNA helicase. Unlike a previously reported DEAD box RNA helicase (LOW EXPRESSION OF OSMOTICALLY RESPONSIVE GENES4 [LOS4]) that regulates mRNA export, RCF1 does not play a role in mRNA export. Instead, RCF1 functions to maintain proper splicing of pre-mRNAs; many cold-responsive genes are misspliced in rcf1-1 mutant plants under cold stress. Functional characterization of four genes (PSEUDO-RESPONSE REGULATOR5 [PRR5], SHAGGY-LIKE SERINE/THREONINE KINASE12 [SK12], MYB FAMILY TRANSCRIPTION FACTOR CIRCADIAN1 [CIR1], and SPFH/PHB DOMAIN-CONTAINING MEMBRANE-ASSOCIATED PROTEIN [SPFH]) that are misspliced in rcf1-1 revealed that these genes are cold-inducible positive (CIR1 and SPFH) and negative (PRR5 and SK12) regulators of cold-responsive genes and cold tolerance. Together, our results suggest that the cold-inducible RNA helicase RCF1 is essential for pre-mRNA splicing and is important for cold-responsive gene regulation and cold tolerance in plants.
DOI:10.1105/tpc.113.111922URLPMID:23771891 [本文引用: 2]
Pre-mRNA splicing is a critical process in gene expression in eukaryotic cells. A multitude of proteins are known to be involved in pre-mRNA splicing in plants; however, the physiological roles of only some of these have been examined. Here, we investigated the developmental roles of a pre-mRNA splicing factor by analyzing root initiation defective1-1 (rid1-1), an Arabidopsis thaliana mutant previously shown to have severe defects in hypocotyl dedifferentiation and de novo meristem formation in tissue culture under high-temperature conditions. Phenotypic analysis in planta indicated that RID1 is differentially required during development and has roles in processes such as meristem maintenance, leaf morphogenesis, and root morphogenesis. RID1 was identified as encoding a DEAH-box RNA helicase implicated in pre-mRNA splicing. Transient expression analysis using intron-containing reporter genes showed that pre-mRNA splicing efficiency was affected by the rid1 mutation, which supported the presumed function of RID1 in pre-mRNA splicing. Our results collectively suggest that robust levels of pre-mRNA splicing are critical for several specific aspects of plant development.
DOI:10.1371/journal.pgen.1005844URLPMID:26848586 [本文引用: 2]
Plants have evolved a considerable number of intrinsic tolerance strategies to acclimate to ambient temperature increase. However, their molecular mechanisms remain largely obscure. Here we report a DEAD-box RNA helicase, TOGR1 (Thermotolerant Growth Required1), prerequisite for rice growth themotolerance. Regulated by both temperature and the circadian clock, its expression is tightly coupled to daily temperature fluctuations and its helicase activities directly promoted by temperature increase. Located in the nucleolus and associated with the small subunit (SSU) pre-rRNA processome, TOGR1 maintains a normal rRNA homeostasis at high temperature. Natural variation in its transcript level is positively correlated with plant height and its overexpression significantly improves rice growth under hot conditions. Our findings reveal a novel molecular mechanism of RNA helicase as a key chaperone for rRNA homeostasis required for rice thermotolerant growth and provide a potential strategy to breed heat-tolerant crops by modulating the expression of TOGR1 and its orthologs.
DOI:10.1111/tpj.12371URLPMID:4350433 [本文引用: 1]
TheArabidopsis thalianaT-DNA insertion mutantrh57-1exhibited hypersensitivity to glucose (Glc) and abscisic acid (ABA). The other tworh57mutants also showed Glc hypersensitivity similar torh57-1, strongly suggesting that the Glc-hypersensitive feature of these mutants results from mutation ofAtRH57.rh57-1andrh57-3displayed severely impaired seedling growth when grown in Glc concentrations higher than 3%. The gene,AtRH57(At3g09720), was expressed in all Arabidopsis organs and its transcript was significantly induced by ABA, high Glc and salt. The newAtRH57belongs to class II DEAD-box RNA helicase gene family. Transient expression ofAtRH57-EGFP(enhanced green fluorescent protein) in onion cells indicated that AtRH57 was localized in the nucleus and nucleolus. Purified AtRH57-His protein was shown to unwind double-stranded RNA independent of ATPin vitro. The ABA biosynthesis inhibitor fluridone profoundly redeemed seedling growth arrest mediated by sugar.rh57-1showed increased ABA levels when exposed to high Glc. Quantitative real time polymerase chain reaction analysis showed thatAtRH57acts in a signaling network downstream ofHXK1. A feedback inhibition of ABA accumulation mediated by AtRH57 exists within the sugar-mediated ABA signaling. AtRH57 mutation and high Glc conditions additively caused a severe defect in small ribosomal subunit formation. The accumulation of abnormal pre-rRNA and resistance to protein synthesis-related antibiotics were observed inrh57mutants and in the wild-type Col-0 under high Glc conditions. These results suggested that AtRH57 plays an important role in rRNA biogenesis in Arabidopsis and participates in response to sugar involving Glc- and ABA signaling during germination and seedling growth.
DOI:10.1093/pcp/pcq045URLPMID:20378763 [本文引用: 1]
DEAD-box RNA helicases are involved in RNA metabolism, including pre-mRNA splicing, ribosome biogenesis, RNA decay and gene expression. In this study, we identified a homolog of the RH36 gene, AtRH36, which encodes a DEAD-box protein in Arabidopsis thaliana. The gene was expressed ubiquitously throughout the plant. The AtRH36 fused to green fluorescent protein was localized in the nucleus. Homozygosity for the Arabidopsis atrh36 mutants, atrh36-1 and atrh36-2, could not be obtained. Progeny of selfed Arabidopsis atrh36 heterozygote plants were obtained at a heterozygote to wild-type ratio of 1 : 1, which suggested that the AtRH36 gene was involved in gametogenesis. Therefore, we performed a reciprocal cross to determine whether AtRH36 was involved in female gametophyte development. Female gametogenesis was delayed in atrh36-1, and asynchronous development of the female gametophytes was found within a single pistil. Knock-down of AtRH36 gave a pleiotropic phenotype and led to the accumulation of unprocessed 18S pre-rRNA. These results suggest that AtRH36 is essential for mitotic division during female gametogenesis and plays an important role in rRNA biogenesis in Arabidopsis.
DOI:10.1007/s11103-010-9659-7URLPMID:20607362 [本文引用: 1]
DEAD-box proteins comprise a large family and function in RNA metabolism. Although DEAD-box proteins are highly conserved among eukaryotes, little is known about the role of DEAD-box proteins in rice. In this study, we identified a rice DEAD-box protein, OsRH36, and demonstrated that OsRH36 and AtRH36 are functional orthologs. OsRH36 was expressed ubiquitously throughout the plant and the OsRH36-GFP fusion protein was localized to the nucleus and nucleolus. Furthermore, functional complementation tests among three homologous DEAD-box protein genes indicated that OsRH36 can restore segregation distortion in the Arabidopsis atrh36 - 1 mutant, but cannot complement the lethal phenotype in the yeast dbp8p mutant. Moreover, mutation of osrh36 also led to defective transmission of the osrh36 allele, as shown for the atrh36 mutants of Arabidopsis . Previously, we have shown that AtRH36 is involved at an early stage of rRNA maturation and is required for synchronous development of female gametophytes in Arabidopsis . Therefore, we suggest that OsRH36 is required for rRNA biogenesis and megagametogenesis in rice, consistent with the function of AtRH36 in Arabidopsis .
DOI:10.1186/s12870-016-0769-5URL [本文引用: 1]
DOI:10.1093/pcp/pcv188URLPMID:26637537 [本文引用: 1]
DEAD-box RNA helicases belong to an RNA helicase family that plays specific roles in various RNA processes, including , mRNA splicing, RNA export, mRNA and RNA decay. This study investigated a DEAD-box RNA helicase, AtRH7/, in . Expression of AtRH7/was ubiquitous; however, the levels of mRNA accumulation were increased in regions and were induced by cold stress. The phenotypes of two allelic AtRH7/-knockout mutants, atrh7-2 and atrh7-3, resembled auxin-related developmental that were exhibited in several ribosomal mutants, and were more severe under cold stress. Northern blot and circular RT-PCR analyses indicated that unprocessed 18S pre-rRNAs accumulated in the atrh7 mutants. The atrh7 mutants were hyposensitive to the antibiotic , which targets , suggesting that AtRH7 was also involved in . In addition, the atrh7-2 and atrh7-3 mutants displayed cold and decreased expression of , and , which might be responsible for the cold intolerance. The present study indicated that AtRH7 participates in rRNA biogenesis and is also involved in plant development and cold tolerance in .
DOI:10.1093/pcp/pcn125URLPMID:18725370 [本文引用: 2]
DEAD-box RNA helicases have been implicated to have a function during stress adaptation processes, but their functional roles in plant stress responses remain to be clearly elucidated. Here, we assessed the expression patterns and functional roles of two RNA helicases, AtRH9 and AtRH25, in Arabidopsis thaliana under abiotic stress conditions. The transcript levels of AtRH9 and AtRH25 were up-regulated markedly in response to cold stress, whereas their transcript levels were down-regulated by salt or drought stress. Phenotypic analysis of the transgenic plants and T-DNA-tagged mutants showed that the constitutive overexpression of AtRH9 or AtRH25 resulted in the retarded seed germination of Arabidopsis plants under salt stress conditions. AtRH25, but not AtRH9, enhanced freezing tolerance in Arabidopsis plants. Both AtRH9 and AtRH25 complemented the cold-sensitive phenotype of BX04 Escherichia coli mutant cells, but AtRH25 had much more prominent complementation ability than AtRH9. An in vitro nucleic acid binding assay showed that AtRH9 binds equally to all homoribopolymers, whereas AtRH25 binds preferentially to poly(G). Taken together, these results demonstrate that AtRH9 and AtRH25 impact on the seed germination of Arabidopsis plants under salt stress conditions, and suggest that the difference in cold tolerance capability between AtRH9 and AtRH25 arises from their different nucleic acid-binding properties.
DOI:10.1111/pce.13200URL [本文引用: 1]
Plants are frequently exposed to numerous environmental stresses such as dehydration and high salinity, and have developed elaborate mechanisms to counteract the deleterious effects of stress. The phytohormone abscisic acid (ABA) plays a critical role as an integrator of plant responses to water-limited condition to activate ABA signal transduction pathway. Although perception of ABA has been... [Show full abstract]
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DOI:10.4161/rna.7.6.13615URLPMID:21045544 [本文引用: 1]
Many non-coding RNAs fold into complex three-dimensional structures, yet the self-assembly of RNA structure is hampered by mispairing, weak tertiary interactions, electrostatic barriers, and the frequent requirement that the 5’ and 3’ ends of the transcript interact. This rugged free energy landscape for RNA folding means that some RNA molecules in a population rapidly form their native structure, while many others become kinetically trapped in misfolded conformations. Transient binding of RNA chaperone proteins destabilize misfolded intermediates and lower the transition states between conformations, producing a smoother landscape that increases the rate of folding and the probability that a molecule will find the native structure. DEAD-box proteins couple the chemical potential of ATP hydrolysis with repetitive cycles of RNA binding and release, expanding the range of conditions under which they can refold RNA structures.
DOI:10.1093/pcp/pcj047URLPMID:16608866 [本文引用: 1]
The mechanistic role of a glycine-rich RNA-binding protein designated atRZ-1a that contributes to enhance cold tolerance in Arabidopsis was investigated. Overexpression of atRZ-1a did not affect the expression of various cold-responsive genes such as COR6.6, COR15a, COR47, RD29A, RD29B and LTI29. Proteome analyses revealed that overexpression of atRZ-1a modulated the expression of several stress-responsive genes, and the transcript levels and RNA stability of these target genes were not affected by atRZ-1a. atRZ-1a successfully complements the cold sensitivity of Escherichia coli lacking four cold shock proteins. These results strongly suggest that atRZ-1a plays a role as an RNA chaperone during the cold adaptation process.
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DOI:10.1177/146642400112100214URLPMID:3134357 [本文引用: 1]
Although glycine-rich RNA-binding proteins (GRPs) have been determined to function as RNA chaperones during the cold adaptation process, the structural features relevant to this RNA chaperone activity remain largely unknown. To uncover which structural determinants are necessary for RNA chaperone activity of GRPs, the importance of the N-terminal RNA recognition motif (RRM) and the C-terminal glycine-rich domains of two Arabidopsis thaliana GRPs (AtGRP4 harbouring no RNA chaperone activity and AtGRP7 harbouring RNA chaperone activity) was assessed via domain swapping and mutation analyses. The results of domain swapping and deletion experiments showed that the domain sequences encompassing the N-terminal RRM of GRPs were found to be crucial to the ability to complement cold-sensitive Escherichia coli mutant cells under cold stress, RNA melting ability, and freezing tolerance ability in the grp7 loss-of-function Arabidopsis mutant. In particular, the N-terminal 24 amino acid extension of AtGRP4 impedes the RNA chaperone activity. Collectively, these results reveal that domain sequences and overall folding of GRPs governed by a specific modular arrangement of RRM and glycine-rich sequences are critical to the RNA chaperone activity of GRPs during the cold adaptation process in cells.
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DOI:10.1016/j.jplph.2015.10.003URLPMID:26513458 [本文引用: 1]
Although the roles of S1 domain-containing proteins have been characterized in diverse cellular processes in the cytoplasm, the functional roles of a majority of S1 domain-containing proteins targeted to the chloroplast are largely unknown. Here, we characterized the function of a nuclear-encoded chloroplast-targeted protein harboring two S1 domains, designated SRRP1 (for S1 RNA-binding ribosomal protein 1), inArabidopsis thaliana. Subcellular localization analysis of SRRP1-GFP fusion proteins revealed that SRRP1 is localized to the chloroplast. The T-DNA tagged loss-of-functionsrrp1mutants displayed poorer seedling growth and less cotyledon greening than the wild-type plants on MS medium supplemented with abscisic acid (ABA), suggesting that SRRP1 plays a role in seedling growth in the presence of ABA. Splicing of thetrnLintron and processing of 5S rRNA in chloroplasts were altered in the mutant plants. Importantly, SRRP1 complemented the growth-defective phenotypes of an RNA chaperone-deficientEscherichia colimutant at low temperatures and had nucleic acid-melting ability, indicating that SRRP1 possesses RNA chaperone activity. Taken together, these results suggest that SRRP1, the chloroplast-localized S1 domain-containing protein, harboring RNA chaperone activity affects the splicing and processing of chloroplast transcripts and plays a role inArabidopsisseedling growth in the presence of ABA.
DOI:10.1111/tpj.12889URLPMID:26031782 [本文引用: 1]
Summary Despite the fact that a variety of nuclear-encoded RNA-binding proteins (RBPs) are targeted to the chloroplast and play essential roles during post-transcriptional RNA metabolism in the chloroplast, the physiological roles of the majority of chloroplast-targeted RBPs remain elusive. Here, we investigated the functional role of a nuclear-encoded S1 domain-containing RBP, designated SDP, in the growth and development of Arabidopsis thaliana . Confocal analysis of the SDP-green fluorescent protein revealed that SDP was localized to the chloroplast. The loss-of-function sdp mutant displayed retarded seed germination and pale-green phenotypes, and grew smaller than the wild-type plants. Chlorophyll a content and photosynthetic activity of the sdp mutant were much lower than those of wild-type plants, and the structures of the chloroplast and the prolamellar body were abnormal in the sdp mutant. The processing of rRNAs in the chloroplast was defective in the sdp mutant, and SDP was able to bind chloroplast 23S , 16S , 5S and 4.5S rRNAs. Notably, SDP possesses RNA chaperone activity. Transcript levels of the nuclear genes involved in chlorophyll biosynthesis were altered in the sdp mutant. Collectively, these results suggest that chloroplast-targeted SDP harboring RNA chaperone activity affects rRNA processing, chloroplast biogenesis and photosynthetic activity, which is crucial for normal growth of Arabidopsis.
DOI:10.1186/1471-2229-14-98URLPMID:4021458 [本文引用: 1]
Background Although several chloroplast RNA splicing and ribosome maturation (CRM) domain-containing proteins have been characterized for intron splicing and rRNA processing during chloroplast gene expression, the functional role of a majority of CRM domain proteins in plant growth and development as well as chloroplast RNA metabolism remains largely unknown. Here, we characterized the developmental and stress response roles of a nuclear-encoded chloroplast protein harboring a single CRM domain (At4g39040), designated CFM4, in Arabidopsis thaliana. Results Analysis of CFM4-GFP fusion proteins revealed that CFM4 is localized to chloroplasts. The loss-of-function T-DNA insertion mutants for CFM4 (cfm4) displayed retarded growth and delayed senescence, suggesting that CFM4 plays a role in growth and development of plants under normal growth conditions. In addition, cfm4 mutants showed retarded seed germination and seedling growth under stress conditions. No alteration in the splicing patterns of intron-containing chloroplast genes was observed in the mutant plants, but the processing of 16S and 4.5S rRNAs was abnormal in the mutant plants. Importantly, CFM4 was determined to possess RNA chaperone activity. Conclusions These results suggest that the chloroplast-targeted CFM4, one of two Arabidopsis genes encoding a single CRM domain-containing protein, harbors RNA chaperone activity and plays a role in the Arabidopsis growth and stress response by affecting rRNA processing in chloroplasts.
