Determining the role of nuclear envelope reformation proteins in regulating the cGAS/STING innate immune response in cancer

项目摘要/摘要： 在癌细胞中，自身DNA暴露在胞浆中是由各种基因组不稳定性驱动的，例如 微核、染色质桥和核破裂。这种胞质DNA可以被胞质DNA识别 传感器，如cGAS(环状GMP-AMP合成酶)，它触发下游的先天免疫反应。 有趣而令人困惑的是，cGAS/STING先天免疫通路的激活可以保护或 根据具体情况使肿瘤对免疫治疗干预措施敏感。因此，洞察 CGAS/STING信号在癌症中的调控方式可以为靶向干预提供信息。来源： 癌细胞中的胞浆DNA主要是由有丝分裂缺陷引起的，这种缺陷导致染色体被包围。 在容易破裂的微核中。这些破裂的微核招募cGAs和核膜 改革(NER)因素-如LEM2、CHMP7和BAF-但仍不清楚这些NER如何或是否 影响cGAS/STING信号转导的因素，但在已发表的和我们的初步数据中有新的证据 CGAS/STING信号和NER蛋白之间存在潜在的串扰。这项提案的目标是 是对癌症中对胞浆DNA的先天性免疫反应的调节提供关键的见解 细胞受核膜重塑因子的影响。为了实现这个目标，我将使用转基因鲱鱼 睾丸(HT)DNA和DNA包被珠作为胞质DNA的模型，因为这可以更容易 与随机形成的微核相比，只有部分微核是不稳定的，容易 破裂。在这个模型中，我将使用CRISPR/Cas9基因编辑和生长素诱导降解(AID)条件 降解系统探讨NER因子在cGAS/STING信号通路中的作用 DNA和DNA珠子。这项提议将涉及细胞生物学和先天免疫的基本方面。 这一信号将有助于阐明癌症的免疫治疗靶点。

PROJECT SUMMARY/ABSTRACT: In cancer cells, exposure of self DNA to the cytosol is driven by a variety of genomic instabilities such as micronuclei, chromatin bridges, and nuclear ruptures. This cytosolic DNA can be recognized by cytosolic DNA sensors such as cGAS (cyclic GMP-AMP synthase), which triggers a downstream innate immune response. Interestingly and confoundingly, the activation of the cGAS/STING innate immune pathway can protect or sensitize tumors to immunotherapeutic interventions depending on the specific context. Therefore, insight into the ways in which cGAS/STING signaling is regulated in cancer can inform targeted intervention. Sources of cytosolic DNA in cancer cells arise primarily from defects in mitosis that lead to the enclosure of chromosomes in micronuclei that are prone to rupture. These ruptured micronuclei recruit cGAS and nuclear envelope reformation (NER) factors—such as LEM2, CHMP7, and BAF—but it remains unknown how, or if, these NER factors impact cGAS/STING signaling but there is emerging evidence in published and in our preliminary data that there is potential crosstalk between cGAS/STING signaling and NER proteins. The goal of this proposal is to provide key insights into the regulation of the innate immune response to cytosolic DNA in cancer cells by nuclear envelope reformation factors. In order to achieve this goal, I will use transfected herring testes (HT) DNA and transfected DNA-coated beads as models for cytosolic DNA as this can be more readily controlled compared to the stochastic formation of micronuclei, only some of which are unstable and prone to rupture. With this model, I will use CRISPR/Cas9 gene-editing and the auxin-inducible-degron (AID) conditional degradation system to probe the roles of NER factors in cGAS/STING signaling in response to transfected HT DNA and DNA beads. This proposal will address fundamental aspects of cell biology and innate immune signaling that will shed light on immunotherapeutic targets for cancer.