Role of Chromatin Bridges in Activating Innate Immune Signaling following Failed Mitosis

项目总结/摘要 由细胞分裂错误产生的异常染色质结构可导致细胞死亡， 不稳定性，并潜在地诱导炎症信号传导。染色质桥（CB）是两个 在癌症和癌症相关疾病的细胞中经常观察到的异常结构类型 Blooms Syndrome（BS）异常的染色质结构可以使基因组自身DNA暴露于 激活抗病毒炎症信号的细胞质。细胞质自身DNA（cyDNA）触发1型 干扰素（IFN）信号通过多种先天免疫途径，其中cGAS-STING轴是最重要的 突出。然而，仅研究了微核诱导IFN信号传导的潜力。之前 报告表明微核自发破裂并产生cyDNA，然后cyDNA激活cGAS。 令人惊讶的是，我已经确定CB，而不是微核，负责cGAS激活和IFN诱导 有丝分裂失败后我建议研究cGAS的分子和物理机制 CB激活。我将使用药物诱导的有丝分裂失败或BS的细胞模型来研究CB。我会调查 cGAS激活CB如何由细胞分裂中的错误产生，CB如何激活cGAS以及cGAS是否 在BS的细胞模型中产生激活CB。我假设炎症性CB是由 未解决的连接或部分连接，并通过张力依赖性机制激活cGAS。 为了验证这一假设，我首先将分析由药物诱导的有丝分裂失败产生的CB，以确定它们的 形成机制然后，我将通过替代方法产生CB，例如拓扑异构酶抑制， 以确定这些CB是否类似地激活cGAS。在我的第二个目标，我将调查是否肌动蛋白- cGAS激活需要CB上的介导张力。我将检验我的假设， 通过挤出染色质结合的蛋白质促进cGAS活化。在目标3中，我将研究细胞 BS模型BS是一种以染色体不稳定为特征的遗传性癌症易感综合征 频繁的CB BS是由BLM解旋酶的功能缺失突变和具有缺陷的 BLM解旋酶在分裂后期表现出大量未分辨的姐妹染色单体连锁。我会学习 这些连接是否变成cGAS激活CB，如果这是通过张力依赖性的 目标2中确定的机制。Mitchison实验室为我提供了一个很好的空间来探索 有丝分裂机制和先天免疫信号。我们在生物学上有了重大发现， 哺乳动物细胞分裂和抗有丝分裂化疗的机制。我提议参加这个传统 通过研究有丝分裂失败的炎症后果及其在Blooms病理学中的潜在作用， 综合征

PROJECT SUMMARY/ABSTRACT Abnormal chromatin structures produced by errors in cell division can cause cell death, propagate genetic instability and, potentially, induce inflammatory signaling. Micronuclei and chromatin bridges (CBs) are two types of abnormal structures which are frequently observed in cells from cancers and cancer-related diseases such as Blooms Syndrome (BS). Aberrant chromatin structures can expose genomic self-DNA to the cytoplasm which activates anti-viral inflammatory signaling. Cytoplasmic self-DNA (cyDNA) triggers type-1 interferon (IFN) signaling through multiple innate immune pathways, of which the cGAS-STING axis is most prominent. However, only micronuclei have been studied for their potential to induce IFN signaling. Prior reports suggested that micronuclei spontaneously rupture and produce cyDNA which then activates cGAS. Surprisingly, I have identified that CBs, not micronuclei, are responsible for cGAS activation and IFN induction after failed mitosis. I propose to investigate the molecular and physical mechanisms underlying cGAS activation by CBs. I will use drug-induced mitotic failure or cellular models of BS to study CBs. I will investigate how cGAS-activating CBs arise from errors in cell division, how CBs activate cGAS and whether cGAS- activating CBs are generated in cellular models of BS. I hypothesize that inflammatory CBs are formed by unresolved catenations or merotelic attachments and activate cGAS through a tension-dependent mechanism. To test this hypothesis, I first will analyze CBs produced by drug-induced mitotic failure to determine their formation mechanism. I then will generate CBs through alternative methods, such as topoisomerase inhibition, to determine whether these CBs similarly activate cGAS. In my second aim I will investigate whether actin- mediated tension across the CB is required for cGAS activation. I will test my hypothesis that tension on CBs facilitates cGAS activation through the extrusion of chromatin-bound proteins. In aim 3 I will study cellular models of BS. BS is an inherited cancer pre-disposition syndrome characterized by chromosomal instability and frequent CBs. BS is caused by loss of function mutations in the BLM helicase and cells with a defective BLM helicase exhibit a high number of unresolved sister chromatid catenations during anaphase. I will study whether these catenations turn into cGAS-activating CBs, and if this occurs through the tension-dependent mechanism identified in aim 2. The Mitchison lab provides me an excellent space to explore the nexus of mitotic mechanisms and innate immune signaling. We have made important discoveries in the biology of mammalian cell division and the mechanisms of anti-mitotic chemotherapy. I propose to partake in this tradition by studying the inflammatory consequences of mitotic failure and its potential role in the pathology of Blooms Syndrome.