The role of PARP10 in alleviating replication stress and promoting cellular proliferation and tumorigenesis

PARP10在缓解复制应激、促进细胞增殖和肿瘤发生中的作用

• 批准号: 10552014
• 负责人: Claudia M Nicolae
• 金额: $ 36.09万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10552014
• 关键词: ADP Ribose Transferases; Address; Adenosine Diphosphate Ribose; Apoptosis; Biochemical; Biological; Biological Assay; Breast; Bypass; CRISPR/Cas technology; Cell Cycle Progression; Cell Line; Cell Proliferation; Cell physiology; Cells; Cellular biology; Chromosome Fragile Sites; Clinic; Coupled; DNA; DNA Biochemistry; DNA Repair; DNA Structure; DNA biosynthesis; DNA lesion; DNA replication fork; DNA sequencing; DNA-Directed DNA Polymerase; Down-Regulation; Enzymes; Epithelium; Exposure to; Family; Fiber; Genes; Genetic Models; Genetic Transcription; Genome Stability; Genomic DNA; Genomic Instability; Genomics; Growth; Hela Cells; Human; Immunocompromised Host; In Vitro; Knock-out; Malignant Neoplasms; Measures; Mediating; Mitochondria; Modeling; Molecular; Mus; Mutagenesis; Mutation; Normal Cell; Nucleic Acids; Oncogenes; Oncogenic; Ovarian; Pathway interactions; Phenotype; Poly(ADP-ribose) Polymerases; Polymerase; Post-Translational Protein Processing; Process; Proliferating; Property; Proteins; Resistance; Role; S phase; Signal Transduction; Structure; Testing; Transgenic Mice; Tumor Promotion; Ubiquitination; Validation; brca gene; cancer cell; carcinogenesis; cell motility; design; genetic manipulation; genome editing; genomic tools; in vivo; inhibitor; member; mouse genetics; mouse model; mutant; next generation; next generation sequencing; novel; overexpression; oxidation; personalized cancer therapy; recruit; replication stress; targeted cancer therapy; transcriptome sequencing; transcriptomics; tumor; tumor growth; tumor xenograft; tumorigenesis

Project Summary Identification of molecular pathways that are preferentially employed and relied upon by cancer cells compared to normal cells is key to designing novel personalized cancer therapies. PARP10 is a poorly characterized member of the PARP family. We previously showed that PARP10 promotes translesion synthesis (TLS)- mediated bypass of DNA lesions during DNA replication, thereby alleviating replication stress. More recently, we also showed that PARP10 is a novel oncogene. We found that the PARP10 gene is amplified and/or overexpressed in a large number of tumors including breast and ovarian, with very few observed occurrences of downregulation or loss. We found that the PARP10 gene is amplified and/or overexpressed in a large proportion of human tumors including breast and ovarian, with almost no occurrences of downregulation or loss. Moreover, we found that PARP10 overexpression in, non-transformed human epithelial RPE1 cells results in enhanced proliferation, resistance to replication stress, and increased xenograft tumor formation in immunocompromised mice. The opposing phenotypes were found upon knockout of PARP10 in cancer HeLa cells. These findings suggest that PARP10 is a putative oncogene and its expression promotes tumor formation and growth. Mutagenic TLS has been previously proposed to promote transformation by both allowing hyper-proliferation and inducing genomic instability. Thus, we hypothesize that PARP10 expression suppresses replication stress through TLS-mediated bypass of replication arresting structures, thereby allowing hyper-proliferation of cancer cells. We propose here to directly test this, in three specific aims which address the hypothesis at three different levels: Aim 1 will investigate the mechanism employed by PARP10 to modulate PCNA-dependent TLS at the molecular level, using biochemical and cellular localization and interaction assays. Aim 2 will functionally test the impact of this mechanism of cellular processes including genomic stability and DNA replication. Aim 3 will employ a mouse genetic model to unambiguously investigate if Parp10 expression induces tumor formation or promotes tumor growth. Using state-of-the-art cellular, molecular and genomic tools (including: CRISPR/Cas9-mediated genome editing; molecular DNA fiber combing to measure fork stability; next generation sequencing approaches to measure mutagenesis and mutation burden) we will investigate here the molecular mechanisms underlying this novel oncogenic function of PARP10. This may eventually result in validation of a new target for cancer therapy.

项目摘要 比较鉴定癌细胞优先使用和依赖的分子途径 对正常细胞是设计新型个性化癌症疗法的关键。 PARP10的特征很差 PARP家族的成员。我们先前表明PARP10促进了跨跨合成（TLS） - 在DNA复制过程中介导的DNA病变旁路，从而减轻了复制应力。最近， 我们还表明，PARP10是一种新颖的癌基因。我们发现PARP10基因被放大和/或 在包括乳房和卵巢在内的大量肿瘤中过表达，很少观察到发生 下调或损失。我们发现PARP10基因在大的 包括乳腺和卵巢在内的人类肿瘤的比例，几乎没有下调或 损失。此外，我们发现非转化的人上皮RPE1细胞中的PARP10过表达 导致增殖增强，对复制应力的抗性以及增加的异种移植肿瘤形成 免疫功能低下的小鼠。在癌症HeLa中敲除PARP10时发现了相反的表型 细胞。这些发现表明PARP10是一种假定的致癌基因，其表达促进了肿瘤 形成和成长。先前已提出了诱变的TLS来促进两者的转化 允许过度增殖并诱导基因组不稳定性。因此，我们假设PARP10表达 通过TLS介导的复制逮捕结构的旁路抑制复制应力，从而抑制重复应力 允许癌细胞过度散热。我们在这里建议直接测试这一点，这是三个特定的目标 解决三个不同层次的假设：AIM 1将研究PARP10所采用的机制 使用生化和细胞定位调节分子水平的PCNA依赖性TLS 互动测定。 AIM 2将在功能上测试这种细胞过程机制的影响，包括 基因组稳定性和DNA复制。 AIM 3将采用小鼠遗传模型进行明确研究 如果PARP10表达诱导肿瘤形成或促进肿瘤生长。使用最先进的细胞， 分子和基因组工具（包括：CRISPR/CAS9介导的基因组编辑；分子DNA纤维 梳理以测量叉子稳定性；下一代测序方法测量诱变和 突变负担）我们将在这里研究这种新型致癌功能的分子机制 PARP10。这最终可能导致验证癌症治疗的新靶标。