Stabilization of Heterochromatin by CLOCK Counteracts hMSC Senescence and Articular Degeneration

Accumulating evidence indicates the close association between the circadian clock and the aging process. However, it is unclear whether and how the circadian clock proteins regulate stem cell aging. Here, we identified a noncanonical transcriptional activator-independent role of CLOCK, a core component of the molecular circadian clock machinery, in counteracting human mesenchymal stem cell (hMSC) senescence. CLOCK expression was decreased during hMSC aging. In addition, CLOCK deficiency accelerated hMSC senescence, whereas CLOCK overexpression attenuated physiological and pathological hMSC senescence. Mechanistic studies revealed that CLOCK formed complexes with nuclear lamina proteins and KAP1, thus maintaining heterochromatin architecture and stabilizing genomic repetitive sequences. Finally, gene therapy with lentiviral vectors encoding CLOCK promoted cartilage regeneration and attenuated age-related osteoarthritis in mice. These findings reveal a novel role of CLOCK independent of its transcriptional activation activity in the stabilization of heterochromatin and alleviation of hMSC senescence, providing a potential therapeutic target for treating aging-associated articular degeneration. Overall design: Multi-omics sequencing of WT and CLOCK defecient hMSCs.

越来越多的研究证据表明，生物钟 (circadian clock) 与衰老进程密切相关。但目前学界尚未明确生物钟蛋白能否调控干细胞衰老，以及其具体的调控机制。本研究首次揭示了分子生物钟核心组件CLOCK的非经典转录激活非依赖性功能：其可拮抗人类间充质干细胞 (human mesenchymal stem cell, hMSC) 的衰老。在hMSC衰老过程中，CLOCK的表达水平显著下调。进一步实验显示，CLOCK缺陷会加速hMSC衰老，而过表达CLOCK则可减轻生理及病理性hMSC衰老。机制研究表明，CLOCK可与核纤层蛋白及KAP1形成复合物，进而维持异染色质结构稳定，并保障基因组重复序列的稳定性。最后，通过编码CLOCK的慢病毒载体进行基因治疗，可促进小鼠软骨再生，并减轻年龄相关性骨关节炎。本研究揭示了CLOCK不依赖转录激活活性的全新功能：稳定异染色质结构并缓解hMSC衰老，为治疗衰老相关关节退行性病变提供了潜在的治疗靶点。整体实验设计：对野生型 (wild-type, WT) 及CLOCK缺陷型hMSC开展多组学测序。