Immune control and genomic instability at micronuclei

项目摘要 染色体不稳定性（CIN）是癌症的一个标志，其特征是染色体错配率高， 在细胞分裂期间进行。CIN可产生称为微核的核畸变， 染色体片段在分裂后期滞后，不能与主染色质团结合，形成原始染色体。 核细胞核募集核膜，但结构缺陷导致经常破裂， 区室化以及蛋白质和小泡与细胞质的不受调节的交换。米- 核被膜破裂引起广泛的功能障碍，并与广泛的DNA损伤有关， 基因组重排，包括成簇突变现象，如chromothripsis和kataegis， 这在癌症基因组中很常见。破裂的微核也可以激活促炎性细胞因子， ry cGAS-STING途径，其在抗肿瘤免疫中起重要作用。这些观察提示 微核可能代表癌症中基因组进化和免疫激活的关键平台。机甲- 驱动微核DNA损伤和免疫激活的微生物知之甚少。实验室发现- 报道了内质网（ER）相关的外切核酸酶TREX 1，它在多种 人类免疫性疾病，包括艾卡迪-古铁雷斯综合征，在微核时累积， 清晰的包膜破裂，切除微核DNA并限制cGAS-STING激活。因此，我们认为， TREX 1在人类健康和疾病的关键途径中占据中心地位，具有不同的作用。因此， 最近，有很强的理由来理解TREX 1活性和与细胞溶质结合的机制， DNA.该实验室的长期目标是确定DNA损伤的机制，聚集性突变- nesis和免疫激活的网站核膜破裂。本建议的具体目标是：（1） 阐明TREX 1结构和功能的机制，2）确定TREX 1如何被募集到微核， （3）分析了微核DNA损伤的途径。每个目标都得到广泛的初步支持， 数据目标1将集中在TREX 1中以前未表征的区域，这对于其去- 分级胞质DNA并抑制cGAS活化。目标2将建立在显示TREX 1 DNA结合 其定位于微核的功能是不确定的，而其与ER的关联是必需的。目标3 将使用一种新的方法来纯化微核，以剖析微核DNA损伤的来源。综合起来看， 这些数据将为癌症基因组进化提供基本的见解，解释以前uncharac- 特征化的TREX 1突变导致Aicardi-Goutières综合征，并可能确定新的策略，以提高抗 肿瘤免疫

PROJECT SUMMARY Chromosomal instability (CIN) is a hallmark of cancer characterized by high rates of chromosome mis-segre- gation during cell division. CIN can generate nuclear aberrations termed micronuclei when a chromosome or chromosome fragment lags during anaphase and fails to join the main chromatin mass that will form the prima- ry nucleus. Micronuclei recruit nuclear envelopes but defects in construction lead to frequent rupturing, loss of compartmentalization, and an unregulated exchange of proteins and small vesicles with the cytoplasm. Mi- cronuclear envelope rupturing causes broad dysfunction and is associated with extensive DNA damage and genomic rearrangements, including clustered mutational phenomena such as chromothripsis and kataegis, which are commonly observed in cancer genomes. Ruptured micronuclei can also activate the pro-inflammato- ry cGAS-STING pathway, which plays essential roles in anti-tumor immunity. These observations suggest that micronuclei may represent key platforms for genome evolution and immune activation in cancer. The mecha- nisms driving DNA damage and immune activation at micronuclei are poorly understood. The laboratory dis- covered that the endoplasmic reticulum (ER)-associated exonuclease TREX1, which is mutated in a variety of human immune diseases including Aicardi-Goutières Syndrome, accumulates at micronuclei upon micronu- clear envelope rupture where it resects micronuclear DNA and limits cGAS-STING activation. Therefore, TREX1 occupies central positions in key pathways with diverse roles in human health and disease. Conse- quently, there is strong rationale to understand mechanisms of TREX1 activity and engagement with cytosolic DNA. The long-term goals of the laboratory are to determine mechanisms of DNA damage, clustered mutage- nesis, and immune activation at sites of nuclear envelope rupture. The specific Aims of this proposal are to 1) Elucidate mechanisms of TREX1 structure and function, 2) Determine how TREX1 is recruited to micronuclei, and 3) Dissect pathways of micronuclear DNA damage. Each objective is supported by extensive preliminary data. Aim 1 will focus on a previously uncharacterized region in TREX1, which is essential for its ability to de- grade cytosolic DNA and inhibit cGAS activation. Aim 2 will build on results showing that TREX1 DNA binding function is dispensable for its localization to micronuclei, while its association with the ER is essential. Aim 3 will use a new method to purify micronuclei to dissect sources of micronuclear DNA damage. Taken together, these data will provide fundamental insights into cancer genome evolution, explain how previously uncharac- terized TREX1 mutations cause Aicardi-Goutières syndrome, and may identify new strategies to improve anti- tumor immunity.