An endogenous PI3K interactome promoting astrocyte-mediated neuroprotection identifies a novel association with RNA-binding protein ZC3H14.
An endogenous PI3K interactome promoting astrocyte-mediated neuroprotection identifies a novel association with RNA-binding protein ZC3H14.
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DOI：
10.1074/jbc.ra120.015389
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发表时间：
2021-01
期刊：
影响因子：
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通讯作者：
Sivak JM
中科院分区：
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作者：
Alqawlaq S;Livne-Bar I;Williams D;D'Ercole J;Leung SW;Chan D;Tuccitto A;Datti A;Wrana JL;Corbett AH;Schmitt-Ulms G;Sivak JM
星形胶质细胞可以通过一系列分泌信号支持神经元的存活，这些信号可以保护神经元免受神经毒性、氧化应激和凋亡级联反应的影响。因此，分析星形胶质细胞分泌组的作用可能为这些神经保护机制提供有价值的见解。以前，我们的特点是一个强大的神经保护活性介导的视网膜星形胶质细胞条件培养基（ACM）对视网膜和皮质神经元的代谢应激模型。然而，神经元细胞中这种复杂活性的分子机制仍不清楚。在这里，激酶抑制剂的化学遗传学筛选揭示了磷酸肌醇3-激酶（PI 3 K）作为转导ACM介导的神经保护作用的中心参与者。为了鉴定有助于保护级联反应的其他蛋白质，从暴露于ACM或对照培养基的神经元细胞中免疫沉淀内源性PI 3 K，然后进行MS/MS蛋白质组学分析。这些数据表明，与PI 3 K共免疫沉淀的蛋白质相对较少，令人惊讶的是，只有5种蛋白质受到ACM信号的调节。这些命中包括预期的PI 3 K相互作用物，例如血小板衍生生长因子受体A（PDGFRA），以及新型RNA结合蛋白相互作用物ZC 3 H14（含锌指CCCH型14）和THOC 1（THO复合蛋白1）。特别是，ZC 3 H14最近已成为一个重要的RNA结合蛋白，在转录后调控中具有多种作用。在验证研究中，我们表明ZC 3 H14的PI 3 K募集对于PDGF诱导的神经保护是必要的，并且这种相互作用存在于原代视网膜神经节细胞中。因此，我们确定了一种新的非细胞自主神经保护信号级联介导的PI 3 K，需要招聘ZC 3 H14，并可能提出一个有前途的策略，以促进星形胶质细胞分泌的促生存信号。
Astrocytes can support neuronal survival through a range of secreted signals that protect against neurotoxicity, oxidative stress, and apoptotic cascades. Thus, analyzing the effects of the astrocyte secretome may provide valuable insight into these neuroprotective mechanisms. Previously, we characterized a potent neuroprotective activity mediated by retinal astrocyte conditioned media (ACM) on retinal and cortical neurons in metabolic stress models. However, the molecular mechanism underlying this complex activity in neuronal cells has remained unclear. Here, a chemical genetics screen of kinase inhibitors revealed phosphoinositide 3-kinase (PI3K) as a central player transducing ACM-mediated neuroprotection. To identify additional proteins contributing to the protective cascade, endogenous PI3K was immunoprecipitated from neuronal cells exposed to ACM or control media, followed by MS/MS proteomic analyses. These data pointed toward a relatively small number of proteins that coimmunoprecipitated with PI3K, and surprisingly only five were regulated by the ACM signal. These hits included expected PI3K interactors, such as the platelet-derived growth factor receptor A (PDGFRA), as well as novel RNA-binding protein interactors ZC3H14 (zinc finger CCCH-type containing 14) and THOC1 (THO complex protein 1). In particular, ZC3H14 has recently emerged as an important RNA-binding protein with multiple roles in posttranscriptional regulation. In validation studies, we show that PI3K recruitment of ZC3H14 is necessary for PDGF-induced neuroprotection and that this interaction is present in primary retinal ganglion cells. Thus, we identified a novel non–cell autonomous neuroprotective signaling cascade mediated through PI3K that requires recruitment of ZC3H14 and may present a promising strategy to promote astrocyte-secreted prosurvival signals.