GW182-Free microRNA Silencing Complex Controls Post-transcriptional Gene Expression during Caenorhabditis elegans Embryogenesis.
GW182-Free microRNA Silencing Complex Controls Post-transcriptional Gene Expression during Caenorhabditis elegans Embryogenesis.
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DOI：
10.1371/journal.pgen.1006484
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发表时间：
2016-12
期刊：
影响因子：
4.5
通讯作者：
Simard MJ
中科院分区：
文献类型：
--
作者：
Jannot G;Michaud P;Quévillon Huberdeau M;Morel-Berryman L;Brackbill JA;Piquet S;McJunkin K;Nakanishi K;Simard MJ
MicroRNAs和ArgAerte形成microRNA诱导的沉默复合体或miRISC，招募GW182，导致mRNA降解和/或翻译抑制。尽管miRISC-GW182相互作用具有明显的保守性和分子意义，但目前尚不清楚miRISC-GW182相互作用是否对动物发育过程中的基因沉默是必不可少的。为了探索这个问题，我们研究了GW182同源基因AIN-1和AIN-2之间的关系和对基因沉默的影响，以及microRNA特异的ArgAerte ALG-1。基于人类ArgAerte结构的同源建模表明，ALG-1具有GW182结合所需的保守的色氨酸结合口袋。我们在体外和体内表明，它们的突变严重改变了与AIN-1和AIN-2的关联。Alg-1色氨酸结合口袋突变动物保留了microRNA结合和处理能力，但缺乏报告沉默活性。有趣的是，ALG-1色氨酸结合口袋突变体在幼虫阶段表现出ALG-1的丢失，但足以挽救胚胎的致死性，表明在这个发育阶段ANS与miRISC的关联是可有可无的。ALG-1色氨酸结合口袋突变体对胚胎mir-35微RNA家族靶点的调控能力进一步证明了AINS在miRNA调控中的可有可无。因此，我们的结果表明，在线虫胚胎发生过程中，microRNA途径可以独立于GW182蛋白发挥作用。动物细胞拥有不同种类的小RNA，能够精确控制基因的表达。其中，microRNAs与ArgAerte蛋白(称为miRISC)形成沉默复合体，通过靶向特定的信使RNA来取消蛋白质的产生。虽然已有共识认为miRISCs有效地介导了基因沉默，但目前仍不清楚它们是否存在于动物中的不同类型。在这里，我们报告了线虫microRNA特异的ArgAerte ALG-1的特定突变，这些突变改变了它与GW182蛋白的同源基因的关联，GW182蛋白是miRISC介导的沉默的重要因素。我们对这个突变体的遗传特征表明，在胚胎发生过程中，部分miRISCs在没有GW182直系物的情况下发挥作用。这些发现表明，在动物发育过程中，存在独特的miRISC，可以以不同的方式调节基因表达。
MicroRNAs and Argonaute form the microRNA induced silencing complex or miRISC that recruits GW182, causing mRNA degradation and/or translational repression. Despite the clear conservation and molecular significance, it is unknown if miRISC-GW182 interaction is essential for gene silencing during animal development. Using Caenorhabditis elegans to explore this question, we examined the relationship and effect on gene silencing between the GW182 orthologs, AIN-1 and AIN-2, and the microRNA-specific Argonaute, ALG-1. Homology modeling based on human Argonaute structures indicated that ALG-1 possesses conserved Tryptophan-binding Pockets required for GW182 binding. We show in vitro and in vivo that their mutations severely altered the association with AIN-1 and AIN-2. ALG-1 tryptophan-binding pockets mutant animals retained microRNA-binding and processing ability, but were deficient in reporter silencing activity. Interestingly, the ALG-1 tryptophan-binding pockets mutant phenocopied the loss of alg-1 in worms during larval stages, yet was sufficient to rescue embryonic lethality, indicating the dispensability of AINs association with the miRISC at this developmental stage. The dispensability of AINs in miRNA regulation is further demonstrated by the capacity of ALG-1 tryptophan-binding pockets mutant to regulate a target of the embryonic mir-35 microRNA family. Thus, our results demonstrate that the microRNA pathway can act independently of GW182 proteins during C. elegans embryogenesis. Animal cells possess different small RNA species capable of precisely controlling the gene expression. Among them, microRNAs form a silencing complex with an Argonaute protein (known as miRISC) that abrogates protein production by targeting specific messenger RNAs. While there is a consensus that miRISCs are effective to mediate gene silencing, it is still unclear if they exist in different types in animals. Here we report specific mutations in the C. elegans microRNA-specific Argonaute ALG-1, which alter its association with the orthologs of GW182 proteins, important factors for miRISC-mediated silencing. Our genetic characterization of this mutant shows that part of miRISCs function without the GW182 orthologs during the embryogenesis. These findings suggest the presence of distinctive miRISC that can regulate gene expression in different ways during animal development.
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DOI：
10.1007/978-1-61779-046-1_16
发表时间：
2011-01-01
期刊：
ARGONAUTE PROTEINS: METHODS AND PROTOCOLS
影响因子：
--
作者：
Jannot, Guillaume;Vasquez-Rifo, Alejandro;Simard, Martin J.
通讯作者：
Simard, Martin J.
影响因子：
11.8
作者：
Grosshans, H;Johnson, T;Slack, FJ
通讯作者：
Slack, FJ
影响因子：
4
作者：
Ambros, Victor
通讯作者：
Ambros, Victor
DOI：
10.15252/embj.201592901
发表时间：
2016-06-01
期刊：
The EMBO journal
影响因子：
--
作者：
Kuzuoğlu-Öztürk D;Bhandari D;Huntzinger E;Fauser M;Helms S;Izaurralde E
通讯作者：
Izaurralde E
影响因子：
56.9
作者：
AMBROS, V;HORVITZ, HR
通讯作者：
HORVITZ, HR