Guo Dongkai, Zhu Yao, Wang Hongfeng, Wang Guanghui, Wang Cheng, Ren Haigang
Laboratory of Clinical Pharmacy, Department of Pharmacy, The Affiliated Suzhou Science and Technology Town Hospital of Nanjing Medical University, Suzhou, Jiangsu 215153, P.R. China.
Laboratory of Molecular Neuropathology, Jiangsu Key Laboratory of Neuropsychiatric Disorders and Department of Pharmacology, College of Pharmaceutical Sciences, Soochow University, Suzhou, Jiangsu 215123, P.R. China.
Exp Ther Med. 2020 Sep;20(3):2639-2648. doi: 10.3892/etm.2020.8988. Epub 2020 Jul 10.
Circadian rhythm serves an essential role in numerous physiological functions. Circadian oscillations are organized by circadian clock components at the molecular level. The precision of the circadian clock is controlled by transcriptional-translational negative feedback loops, as well as post-translational modifications of clock proteins, including ubiquitination; however, the influence of E3 ligases on clock protein ubiquitination requires further investigation. The results of co-immunoprecipitation and immunofluorescent localization, indicated that the endoplasmic reticulum transmembrane E3 ubiquitin ligase HRD1, encoded by the synoviolin 1 gene, interacted with brain and muscle ARNT-like 1 (BMAL1) and enhanced BMAL1 protein ubiquitination. In addition, the results of western blotting and reverse transcription-quantitative PCR suggested that HRD1 promoted K48-associated polyubiquitination of BMAL1 and thus mediated its degradation via the ubiquitin-proteasome system. Furthermore, gene knockdown and gene overexpression assays revealed that HRD1-dependent degradation of BMAL1 protein regulated the expression of BMAL1 target genes and the amplitude of circadian oscillations in mammalian cells. The findings of the current study indicate that HRD1 may influence the regulation of circadian rhythm via modulation of BMAL1 stability.
昼夜节律在众多生理功能中起着至关重要的作用。昼夜振荡在分子水平上由生物钟组件组织而成。生物钟的精确性由转录 - 翻译负反馈环以及包括泛素化在内的生物钟蛋白的翻译后修饰所控制；然而，E3 连接酶对生物钟蛋白泛素化的影响仍需进一步研究。免疫共沉淀和免疫荧光定位结果表明，由滑膜素 1 基因编码的内质网跨膜 E3 泛素连接酶 HRD1 与脑和肌肉芳香烃受体核转运蛋白样 1（BMAL1）相互作用，并增强了 BMAL1 蛋白的泛素化。此外，蛋白质印迹和逆转录 - 定量 PCR 结果表明，HRD1 促进了 BMAL1 的 K48 相关多聚泛素化，从而通过泛素 - 蛋白酶体系统介导其降解。此外，基因敲低和基因过表达实验表明，HRD1 依赖的 BMAL1 蛋白降解调节了 BMAL1 靶基因的表达以及哺乳动物细胞中昼夜振荡的幅度。当前研究结果表明，HRD1 可能通过调节 BMAL1 的稳定性来影响昼夜节律的调控。
