A Recessive Mutant of argonaute1b/gsnl4 Leads to Narrow Leaf, Small Grain Size and Low Seed Setting in Rice
A Recessive Mutant of argonaute1b/gsnl4 Leads to Narrow Leaf, Small Grain Size and Low Seed Setting in Rice
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argonaute1b/gsnl4 隐性突变体导致水稻叶片窄、籽粒小、结实率低
DOI：
10.1016/j.rsci.2021.05.012
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发表时间：
2021-11
期刊：
影响因子：
4.8
通讯作者：
Guo LongBiao
中科院分区：
文献类型：
--
作者：
Song Mengqiu;Shen Lan;Peng Youlin;Hu Haitao;Ding Shilin;Ruan Shuang;Cui Yongtao;Jiang Hongzhen;Zhang Yu;Wang Zhongwei;Gao Zhenyu;Jahan Noushin;Hu XingMing;Qian Qian;Guo LongBiao
利用甲基磺酸乙酯（EMS）诱变粳稻武运粳21号，获得了一个小粒、窄叶、结实率低的突变体gsnl 4。遗传分析表明gsnl 4是一个功能缺失突变体。OsAGO 1b蛋白Piwi结构域中的gsnl 4单碱基突变导致Ser被Asn取代。CRISPR/Cas9介导的OsAGOlb编辑产生了表型上类似gsnl 4的突变体。此外，miRNA-Seq分析表明，与花粉发育、叶片形态和激素激活相关的信号转导途径中的miRNAs转录表达水平在gsnl 4突变体和野生型（WT）植物之间存在显著差异。在gsnl 4突变体中，几种miRNAs下调，并且它们的靶基因上调。gsnl 4突变体根尖中生长素含量下降，大多数生长素相关基因的表达发生改变。综上所述，GSNL 4不仅通过控制细胞的分裂和扩展来调节器官的发育，而且在调节生长素的运输方面也起着重要的作用。ARGONAUTE（AGO）蛋白广泛分布于真核生物中，是RNA诱导沉默复合物（RISC）的核心组分（Baulcombe，2004）。AGO蛋白与小的非编码RNA如siRNA和miRNA结合，通过影响蛋白质合成和RNA稳定性在RNA沉默机制中发挥重要作用。AGO蛋白含有四个保守结构域：N-末端结构域、PAZ结构域、中间结构域（MID）和Piwi结构域（Song等，2004）。先前的研究表明，Piwi结构域是AGO的催化核心，其与核糖核酸酶H（RNase H）以氨基酸四联体Asp-Glu-Asp-His/Asp（DEDH/D）的保守催化中心的形式共享结构相似性（Song等人，2004）。催化中心的功能是切割小RNA的靶序列。大米含有19种AGO蛋白质。其中，OsAGO 7的突变导致叶片向上卷曲，延长叶片直立时间，促进直立叶冠的形成（Shi et al，2007）。OsAGO 2通过DNA甲基化直接调控OsHXK 1的表达，调控花药发育（Zheng et al，2019）。OsAGO 17可能是siRNA途径中的关键蛋白，正调控水稻籽粒大小和重量，促进水稻茎发育（Zhong et al，2020）。OsAGO 1有4个同源物，命名为OsAGO 1a/B/c/d，它们可能是功能冗余的。RNA干扰OsAGO 1 s保守区域导致植物矮化、叶片变窄和卷曲，以及结实率低（Wu et al，2009）。最近，据报道OsAGO 1b是水稻生长和发育的关键调节因子，但不参与叶极性的建立（Li et al，2019）。然而，这些研究都是基于反向遗传学的，通过正向遗传学研究单个OsAGO 1b基因突变对植物器官发育的影响尚不清楚。本研究以粳稻武运粳21为材料，经电磁辐射诱变，获得了一个多效突变体gsnl 4（粒径窄叶4）。gsnl 4突变体表现出窄叶、小颗粒大小、低花粉育性、低结实率和细秆（图1-A至F;图S1）。为了确定GSNL 4基因对各种组织发育的影响，进行了多个器官的组织学分析。成熟期倒二叶片中部横切面观察表明，成熟期叶片的小脉和大脉总数明显减少。
A gsnl4 mutant characterized by small grain size, narrow leaf and low seed-setting rate was obtained by ethyl methane sulfonate (EMS) mutagenesis of a japonica rice variety Wuyunjing 21. Genetic analysis showed that gsnl4 is a loss-of-function mutant. A single-base mutation in gsnl4 resulted in the substitution of Ser to Asn in the Piwi domain of OsAGO1b protein. CRISPR/Cas9-mediated editing of OsAGO1b yielded a mutant phenotypically resembling gsnl4. Furthermore, miRNA-Seq analysis showed that the transcript expression levels of miRNAs in the signal transduction pathways related to pollen development, leaf morphology and hormone activation were significantly different between the gsnl4 mutant and the wild type (WT) plants. Several miRNAs were downregulated, and their target genes were upregulated in gsnl4 mutants. The auxin content in the root tips of the gsnl4 mutant decreased, and the expression of most auxin-related genes was altered. In summary, GSNL4 not only regulates organ development by controlling cell division and expansion, but also plays an important role in regulating auxin transport in rice. ARGONAUTE (AGO) proteins are widely distributed in eukaryotes and are the central components of the RNA-induced silencing complex (RISC)(Baulcombe, 2004). AGO proteins bind to small non-coding RNAs, such as siRNAs and miRNAs, and play an important role in the silencing mechanism of RNAs by affecting protein synthesis and RNA stability. AGO proteins contain four conserved domains: N-terminal domain, PAZ domain, middle domain (MID) and Piwi domain (Song et al, 2004). Previous studies showed that the Piwi domain is the catalytic core of AGO, which shares structural similarity with ribonuclease H (RNase H) in the form of a conserved catalytic center of amino acid quadruple Asp-Glu-Asp-His/Asp (DEDH/D)(Song et al, 2004). The function of the catalytic center is to cut the target sequence of small RNA. Rice contains 19 AGO proteins. Among them, the mutation of OsAGO7 causes the leaves to curl upwards, extends the upright time of leaves, and promotes the formation of an upright leaf crown (Shi et al, 2007). OsAGO2 directly regulates the expression of OsHXK1 through DNA methylation and regulates the development of anthers (Zheng et al, 2019). OsAGO17 may be a key protein in the siRNA pathway, which positively regulates rice grain size and weight, and promotes rice stem development (Zhong et al, 2020). OsAGO1 has four homologs, named OsAGO1a/b/c/d, which may be functionally redundant. RNA interference on the conservative regions of OsAGO1s results in dwarfism of plants, narrowed and curled leaves, as well as low seed-setting rate (Wu et al, 2009). Recently, OsAGO1b was reported to be a key regulator of growth and development in rice but did not participate in the establishment of leaf polarity (Li et al, 2019). However, these studies were based on reverse genetics, and the effects of individual OsAGO1b gene mutation on development of plant organs is still unclear through forward genetics. In this study, a pleiotropic mutant gsnl4 (grain size and narrow leaf 4) was isolated from a japonica rice Wuyunjing 21 after EMS-induced mutagenesis. The gsnl4 mutant showed narrow leaves, small grain size, low pollen fertility, low seedsetting rate and thin culm (Fig. 1-A to-F; Fig. S1). To determine the effect of the GSNL4 gene on the development of various tissues, histological analysis of multiple organs was performed. The transverse sections of the middle part of the second blades from top at the maturity stage showed that the total numbers of small veins and large veins were significantly reduced in …
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影响因子：
56.9
作者：
Song, JJ;Smith, SK;Joshua-Tor, L
通讯作者：
Joshua-Tor, L
DOI：
10.1073/pnas.1817675116
发表时间：
2019-03
影响因子：
11.1
作者：
Shaoyan Zheng;Jing Li;Lu Ma;Hailong Wang;Hai Zhou;Erdong Ni;Dagang Jiang;Zhenlan Liu;C. Zhuan
通讯作者：
Shaoyan Zheng;Jing Li;Lu Ma;Hailong Wang;Hai Zhou;Erdong Ni;Dagang Jiang;Zhenlan Liu;C. Zhuan
影响因子：
7.2
作者：
Nishimura, A;Ito, M;Matsuoka, M
通讯作者：
Matsuoka, M
影响因子：
10.5
作者：
Vaucheret, H;Vazquez, F;Bartel, DP
通讯作者：
Bartel, DP
影响因子：
7.4
作者：
Tsiantis, M;Brown, MIN;Langdale, JA
通讯作者：
Langdale, JA