Oxygen-dependent changes in binding partners and post-translational modifications regulate the abundance and activity of HIF-1α/2α
Oxygen-dependent changes in binding partners and post-translational modifications regulate the abundance and activity of HIF-1α/2α
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DOI：
10.1126/scisignal.abf6685
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发表时间：
2021-07-20
影响因子：
7.3
通讯作者：
Eyers, Claire E.
中科院分区：
文献类型：
--
作者：
Daly, Leonard A.;Brownridge, Philip J.;Eyers, Claire E.
低氧诱导因子（hypoxia-inducible factors，HIFs）介导细胞对低氧环境的适应。与其他转录因子一样，HIF的稳定性和转录活性以及低氧反应受翻译后修饰（PTM）和蛋白质-蛋白质相互作用变化的调节。我们目前对PTM介导的HIF调控的理解主要基于体外蛋白片段研究，这些研究通常在用低氧模拟化合物处理的片段表达细胞中得到验证。在这里，我们使用基于免疫沉淀的质谱来表征在常氧（21%氧气）和低氧（1%氧气）条件下全长HIF-1 α和HIF-2 α的PTM和结合伴侣。缺氧显著改变了HIF α蛋白相互作用网络的复杂性和组成，特别是对于HIF-2 α，两种亚型的缺氧网络都富含线粒体蛋白。此外，两种HIF α同种型都被大量共价修饰。我们确定了类似的40 PTM网站组成的13种不同类型的修改HIF α亚型，包括多个半胱氨酸修饰和一个不寻常的磷酸半胱氨酸。超过80%的PTM是以前未知的，大约一半表现出氧依赖性。我们进一步表征了HIF-1 α中Ser（31）的进化保守磷酸化作为其转录功能的调节因子，并且我们提出了Thr（406）、Thr（528）和Ser（581）在HIF-2 α中的功能作用。这些数据将有助于描绘不同的生理作用，这些密切相关的亚型微调缺氧反应。
Cellular adaptation to low-oxygen environments is mediated in part by the hypoxia-inducible factors (HIFs). Like other transcription factors, the stability and transcriptional activity of HIFs-and consequently, the hypoxic response-are regulated by post-translational modifications (PTMs) and changes in protein-protein interactions. Our current understanding of PTM-mediated regulation of HIFs is primarily based on in vitro protein fragment-based studies typically validated in fragment-expressing cells treated with hypoxia-mimicking compounds. Here, we used immunoprecipitation-based mass spectrometry to characterize the PTMs and binding partners for full-length HIF-1 alpha and HIF-2 alpha under normoxic (21% oxygen) and hypoxic (1% oxygen) conditions. Hypoxia substantially altered the complexity and composition of the HIF alpha protein interaction networks, particularly for HIF-2 alpha, with the hypoxic networks of both isoforms being enriched for mitochondrial proteins. Moreover, both HIF alpha isoforms were heavily covalently modified. We identified similar to 40 PTM sites composed of 13 different types of modification on both HIF alpha isoforms, including multiple cysteine modifications and an unusual phosphocysteine. More than 80% of the PTMs identified were not previously known and about half exhibited oxygen dependency. We further characterized an evolutionarily conserved phosphorylation of Ser(31) in HIF-1 alpha as a regulator of its transcriptional function, and we propose functional roles for Thr(406), Thr(528), and Ser(581) in HIF-2 alpha. These data will help to delineate the different physiological roles of these closely related isoforms in fine-tuning the hypoxic response.