Phosphorylation of the Drosophila melanogaster RNA-binding protein HOW by MAPK/ERK enhances its dimerization and activity.
Phosphorylation of the Drosophila melanogaster RNA-binding protein HOW by MAPK/ERK enhances its dimerization and activity.
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DOI：
10.1371/journal.pgen.1002632
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发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Volk T
中科院分区：
文献类型：
--
作者：
Nir R;Grossman R;Paroush Z;Volk T
黑腹果蝇伸出翅膀（HOW）是一种保守的rna结合蛋白（RBP），属于STAR家族，其最接近的哺乳动物同源物Quaking （QKI）与胚胎发育和神经系统髓鞘形成有关。RBP如何通过控制mRNA水平和多个关键调控基因的剪接谱来调节多种发育过程；然而，在组织中调节其活性的机制尚未阐明。在这里，我们通过显示HOW被MAPK/ERK磷酸化，将受体酪氨酸激酶（RTK）信号传导与STAR蛋白QKI亚家族的调控联系起来。重要的是，我们发现这种修饰促进了HOW二聚化，增强了其结合RNA和调节其水平的能力。利用特异性识别磷酸化的HOW的抗体，我们发现HOW在胚胎肌肉和心脏母细胞中被磷酸化，从而首次记录了完整生物体中STAR蛋白的丝氨酸/苏氨酸（Ser/Thr）磷酸化。我们还发现sallimus/D-titin （sls）基因是how介导的负调控的一个新的肌肉靶点，并进一步表明这种调控是磷酸化依赖的，强调了这种修饰的生理相关性。重要的是，我们证明了在果蝇MAPK的肌肉特异性敲低后，Thr的磷酸化会减少，相应地，Sls在这些肌肉中升高，类似于HOW RNAi的作用。综上所述，我们的研究结果提供了不同HOW激活的连贯机制；MAPK/ erk依赖性的HOW磷酸化促进HOW二聚体的形成，从而增强其控制关键肌肉特异性基因mRNA水平的活性。因此，我们的研究结果在MAPK/ERK信号和发育中的肌肉RNA调节之间架起了桥梁。体肌是具有高度组织化肌体结构的巨大细胞，其形成和维持尚不完全清楚。多种信号在这些细胞中起作用，包括高度保守的MAPK/ERK通路，它通常作为细胞增殖或分化的线索。在这项研究中，我们揭示了MAPK/ERK信号通过控制果蝇rna结合蛋白hold Out wings （HOW）的活性，在mRNA水平上调控多个肌肉基因的作用。具体来说，我们发现HOW被MAPK/ERK磷酸化，这增加了它形成二聚体的能力，增强了它的rna结合能力。我们进一步证明，在胚胎和幼虫肌肉中，HOW被MAPK磷酸化，这一事件对于其调节与脊椎动物titin同源的巨大肌肉基因水平的能力很重要，从而有助于维持肌肉的肌肉结构。重要的是，HOW是哺乳动物Quaking的同源基因，Quaking是胚胎发育和神经系统髓鞘形成的一种必需蛋白，其减少与精神分裂症有关。因此，我们的研究结果提出了MAPK/ERK磷酸化类似地调节其他组织中由保守的Quaking家族蛋白控制的RNA谱的可能性。
Drosophila melanogaster Held Out Wings (HOW) is a conserved RNA–binding protein (RBP) belonging to the STAR family, whose closest mammalian ortholog Quaking (QKI) has been implicated in embryonic development and nervous system myelination. The HOW RBP modulates a variety of developmental processes by controlling mRNA levels and the splicing profile of multiple key regulatory genes; however, mechanisms regulating its activity in tissues have yet to be elucidated. Here, we link receptor tyrosine kinase (RTK) signaling to the regulation of QKI subfamily of STAR proteins, by showing that HOW undergoes phosphorylation by MAPK/ERK. Importantly, we show that this modification facilitates HOW dimerization and potentiates its ability to bind RNA and regulate its levels. Employing an antibody that specifically recognizes phosphorylated HOW, we show that HOW is phosphorylated in embryonic muscles and heart cardioblasts in vivo, thus documenting for the first time Serine/Threonine (Ser/Thr) phosphorylation of a STAR protein in the context of an intact organism. We also identify the sallimus/D-titin (sls) gene as a novel muscle target of HOW–mediated negative regulation and further show that this regulation is phosphorylation-dependent, underscoring the physiological relevance of this modification. Importantly, we demonstrate that HOW Thr phosphorylation is reduced following muscle-specific knock down of Drosophila MAPK rolled and that, correspondingly, Sls is elevated in these muscles, similarly to the HOW RNAi effect. Taken together, our results provide a coherent mechanism of differential HOW activation; MAPK/ERK-dependent phosphorylation of HOW promotes the formation of HOW dimers and thus enhances its activity in controlling mRNA levels of key muscle-specific genes. Hence, our findings bridge between MAPK/ERK signaling and RNA regulation in developing muscles. Somatic muscles are huge cells that feature highly organized sarcomeric architecture, whose formation and maintenance are not fully understood. Multiple signals play a role in these cells, including the highly conserved MAPK/ERK pathway, which often serves as a cue for cellular proliferation or differentiation. In this study, we reveal a role for MAPK/ERK signaling in the regulation of multiple muscle genes at the level of mRNA, through the control of the activity of the Drosophila RNA–binding protein Held Out wings (HOW). Specifically, we show that HOW undergoes phosphorylation by MAPK/ERK, which increases its ability to form dimers and enhances its RNA–binding capacity. We further demonstrate that HOW is phosphorylated in embryonic and larval muscles by MAPK in vivo and that this event is important for its ability to regulate the levels of a giant sarcomeric gene homologous to vertebrate titin, thus contributing to the maintenance of muscle sarcomeric architecture. Importantly, HOW is a close homolog of mammalian Quaking, an essential protein in embryonic development and nervous system myelination, a reduction of which is correlated with Schizophrenia. Thus, our results raise the possibility that MAPK/ERK phosphorylation similarly regulates RNA profiles in other tissues controlled by proteins of the conserved Quaking family.
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