Functional genetic screens for structural regulatory DNA elements driving tumor progression and drug resistance

人类基因组以两种不同的形式携带遗传信息：转录的编码基因和非编码基因以及非转录的调控DNA元件(RDes)。RDES控制基因表达的大小和模式，对生物体的发育是不可或缺的。与此相一致，越来越多的证据表明rDES的变异是肿瘤发病的关键，这表明rDES在健康和疾病的细胞识别中起着至关重要的作用。rDES可细分为转录rDES(t-rDES)和结构rDES(S-rDES)，前者招募转录因子来增强基因，从而影响基因表达，后者维持染色质3D结构，这是t-rDES调控基因所必需的特征。虽然大规模的功能基因筛查极大地促进了我们在研究哺乳动物基因方面的知识，但这种分析rDES的工具还很缺乏。在过去的几年里，CRISPR/CAS系统在基因组编辑技术方面的惊人进步不仅扩大了可用于检查基因的筛选工具箱，而且为研究rDES开辟了新的可能性。最近，AGami小组开发了一种针对t-rDes的CRISPR策略，并显示了其在识别潜在的致癌和肿瘤抑制t-rde方面的可扩展性和有效性。在拟议的项目中，我们将利用这一经验针对S-rDes，并建立功能基因筛查，以确定在细胞增殖和生存、癌症进展和耐药方面的关键角色。此外，我们还将研究潜在的分子机制。因此，我们的结果将提供必要的功能知识，以更好地理解S-rDES在癌症中控制3D染色质结构的作用及其与癌症进展和耐药性的关系。

The human genome carries genetic information in two distinct forms: Transcribed coding and non-coding genes as well as non-transcribed regulatory DNA elements (rDEs). rDEs control the magnitude and pattern of expression of genes and are indispensable for organismal development. In line with this, increasing evidence indicates that variation of rDEs is key to tumor pathogenesis, suggesting that rDEs play crucial roles in cell identity in health and disease.rDEs can be subdivided into transcriptional rDEs (t-rDEs) that recruit transcription factors to enhancers, thereby affecting gene expression, and structural rDEs (s-rDEs) that maintain chromatin 3D structure, a feature essential for regulation of genes by t-rDEs. While large-scale functional genetic screens greatly advanced our knowledge in studying mammalian genes, such tools to analyze rDEs were lacking. In the last few years, spectacular advance in genome editing technologies by the CRISPR/Cas system has not only expanded the available screening toolbox to examine genes, but also opened up novel possibilities for studying rDEs. Recently, the Agami group has developed a CRISPR strategy to target t-rDEs and has shown its scalability and effectivity in identifying potential oncogenic and tumor-suppressive t-rDEs. In the proposed project, we will exploit this experience for targeting s-rDEs, and setup functional genetic screens to identify key players in cell proliferation and survival, cancer progression and resistance to therapy. Moreover, we will investigate underlying molecular mechanisms. Thereby, our results will provide the functional knowledge necessary to better understand the role of s-rDEs in controlling the 3D chromatin structure in cancer and its implication for cancer progression and drug resistance.