Buck Institute for Research on Aging, Novato, CA 94945, USA.
Department of Plant and Microbial Biology, University of California, Berkeley, CA 94720, USA.
G3 (Bethesda). 2023 Jul 5;13(7). doi: 10.1093/g3journal/jkad091.
Cellular senescence is a program of cell cycle arrest, apoptosis resistance, and cytokine release induced by stress exposure in metazoan cells. Landmark studies in laboratory mice have characterized a number of master senescence regulators, including p16INK4a, p21, NF-κB, p53, and C/EBPβ. To discover other molecular players in senescence, we developed a screening approach to harness the evolutionary divergence between mouse species. We found that primary cells from the Mediterranean mouse Mus spretus, when treated with DNA damage to induce senescence, produced less cytokine and had less-active lysosomes than cells from laboratory Mus musculus. We used allele-specific expression profiling to catalog senescence-dependent cis-regulatory variation between the species at thousands of genes. We then tested for correlation between these expression changes and interspecies sequence variants in the binding sites of transcription factors. Among the emergent candidate senescence regulators, we chose a little-studied cell cycle factor, upstream stimulatory factor 2 (USF2), for molecular validation. In acute irradiation experiments, cells lacking USF2 had compromised DNA damage repair and response. Longer-term senescent cultures without USF2 mounted an exaggerated senescence regulatory program-shutting down cell cycle and DNA repair pathways, and turning up cytokine expression, more avidly than wild-type. We interpret these findings under a model of pro-repair, anti-senescence regulatory function by USF2. Our study affords new insights into the mechanisms by which cells commit to senescence, and serves as a validated proof of concept for natural variation-based regulator screens.
细胞衰老（Cellular senescence）是一种细胞周期停滞、凋亡抵抗和细胞因子释放的程序，由多细胞生物细胞暴露于应激时引发。实验室小鼠的标志性研究已经鉴定了许多主要的衰老调控因子，包括 p16INK4a、p21、NF-κB、p53 和 C/EBPβ。为了发现衰老过程中的其他分子参与者，我们开发了一种筛选方法，利用了鼠科物种之间的进化差异。我们发现，用 DNA 损伤处理来自地中海鼠种 Mus spretus 的原代细胞以诱导衰老时，这些细胞产生的细胞因子较少，溶酶体也不如来自实验室鼠种 Mus musculus 的细胞活跃。我们使用等位基因特异性表达谱，在数千个基因中对物种之间衰老相关的顺式调控变异进行了编目。然后，我们测试了这些表达变化与转录因子结合位点中的种间序列变异之间的相关性。在新兴的候选衰老调控因子中，我们选择了一个研究较少的细胞周期因子，上游刺激因子 2（Upstream stimulatory factor 2，USF2），用于分子验证。在急性辐照实验中，缺乏 USF2 的细胞的 DNA 损伤修复和反应受损。没有 USF2 的长期衰老培养物会引发更夸张的衰老调控程序——关闭细胞周期和 DNA 修复途径，并更强烈地表达细胞因子，比野生型更强烈。我们根据 USF2 的修复促进、抗衰老调控功能模型来解释这些发现。我们的研究为细胞衰老的机制提供了新的见解，并为基于自然变异的调控因子筛选提供了经过验证的概念证明。
