Project description:The human A-family DNA polymerase M-NM-8 (Pol q) is a large, multidomain enzyme whose physiological function is still unclear despite its in vitro translesion synthesis capacity in front of DNA damage and its involvement in some features of DNA repair after external stress. Here we present evidence that Pol q holds a novel role in the absence of external stress as a critical determinant of the replication timing program in human cells. Pol q binds to chromatin at early G1 and is required for proper formation of pre-replicative complexe and replication origin activation. Pol q-depleted cells show modified spatial organization of chromatin-loop structures at replication factories. Genome-wide analysis of replication timing shows delayed replication of a part of early replicating domains and advanced replication of a part of late replicating domains following Pol q depletion. Our results identify Pol q as one of the first critical human factors discovered in the replication timing programme. Two-condition experiment, siRNA control vs. siRNA polQ cells. Biological replicates: 2 control replicates, 2 transfected replicates.
Project description:The human A-family DNA polymerase M-NM-8 (Pol q) is a large, multidomain enzyme whose physiological function is still unclear despite its in vitro translesion synthesis capacity in front of DNA damage and its involvement in some features of DNA repair after external stress. Here we present evidence that Pol q holds a novel role in the absence of external stress as a critical determinant of the replication timing program in human cells. Pol q binds to chromatin at early G1 and is required for proper formation of pre-replicative complexe and replication origin activation. Pol q-depleted cells show modified spatial organization of chromatin-loop structures at replication factories. Genome-wide analysis of replication timing shows delayed replication of a part of early replicating domains and advanced replication of a part of late replicating domains following Pol q depletion. Our results identify Pol q as one of the first critical human factors discovered in the replication timing programme. Two-condition experiment, control vs. over expressed polQ cells. Biological replicates: 2 control replicates, 2 transfected replicates.
Project description:<p>BRCA1 mutations are a hallmark of hereditary ovarian cancer, strongly linked to deficiencies in homologous recombination (HR) DNA repair and impaired DNA replication fork protection. However, its roles in cancer progression beyond maintaining genomic integrity remain poorly understood. Through metabolomics approaches, we found BRCA1-deficiency strikingly increased choline metabolism. Loss of BRCA1 promotes choline uptake through upregulating choline transporter-like protein 4 (CTL4). BRCA1 directly binds and recruits EZH2-mediated H3K27Me3 deposition to CTL4 promoter. CTL4 was therefore overexpressed in ovarian cancer tissues with BRCA1 mutations. Furthermore, BRCA1-deficiency significantly promotes ovarian cancer invasion, while inhibition of CTL4 reverses the high metastatic potential of BRCA1-deficient ovarian cancer cells, suggesting the functionality and specificity of CTL4 as a therapeutic target. Additionally, we discovered that phosphocholine, the choline metabolite increased by CTL4 overexpression, interacted with and stabilized the epithelial-to-mesenchymal transition inducer FAM3C in BRCA1-deficient ovarian cancer cells. Importantly, we identified a potent CTL4 inhibitor, DT-13, which significantly reduces choline metabolism and effectively suppresses metastasis in BRCA1-deficient ovarian cancers. Therefore, our study uncovers a mechanism underlying metastasis in BRCA1-deficient cancers and identifies CTL4 as a therapeutic target for metastatic ovarian cancer patients with BRCA1 mutations.</p>
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.