Saudi Cultural Missions Theses & Dissertations
Permanent URI for this communityhttps://drepo.sdl.edu.sa/handle/20.500.14154/10
Browse
2 results
Search Results
Item Restricted PROTEOMICS ANALYSIS OF RNA BINDING PROTEINS IN PROSTATE CANCER CELLS(Clark Atlanta University, 2024-03-26) Dwaed, Abdulrahman; Cinar, BekirRNA-binding proteins (RBPs) govern various RNA-related functions, and dysregulation of RBPs is linked to numerous human diseases, including cancer. RBPs regulate the intricate facets of RNA biogenesis, impacting its metabolism and functional kinetics. This study investigates the biochemical and functional interactions between the RNA-binding protein NPM1 and the Hippo pathway effector of the transcriptional coregulatory protein YAP1. This study also examined the system-wide changes in RBP patterns in prostate cancer cell lines with differential responses to androgen hormone signaling. This study revealed that androgen hormone signaling regulates the protein-protein interaction of YAP1 with NPM1. GST-pulldown assay combined with western blot showed that the proline-rich region and WW/SH3 domain of YAP1 mediated the interaction with NPM1. The proximity ligation assay combined with co-immunoprecipitation further verified the interaction between YAP1 and NPM1 in the androgen-dependent (AD) and androgen-independent (AI) cell lines. Disruption of NPM1 activity by genetic and pharmacological approaches reduced cell growth and cell migration in cultures, likely mediated by the inhibition of YAP1. Moreover, treatment of AD and AI cells with or without androgen resulted in distinct RBP patterns, as demonstrated by enhanced RNA interactome capture assay combined with quantitative mass spectroscopy-based proteomics. Furthermore, the analysis of prostate cancer tissues showed an elevated NPM1 expression that correlated with prostate tumor progression. These findings suggest that altered RBP patterns downstream of modified androgen hormone signaling contribute to tumor progression, and the YAP1-NPM1 axis may constitute a viable drug target in cancer.17 0Item Restricted Validation of Driver Genetic Aberrations in Undifferentiated Pleomorphic Sarcoma: Focus on the Role of YAP1 Utilising a CRISPR/Cas9 Approach.(Saudi Digital Library, 2023-11-21) Alshammari, Ahmad; Sisley, KarenSoft tissue sarcoma (STS) is a rare, diverse group of tumours that are characterised by genetic, pathological and clinical diversity. STS comprise less than 1% of all adult malignancies and consist of more than 100 histological and molecular subtypes. Undifferentiated pleomorphic sarcoma (UPS) is a sarcoma with no specific line of differentiation and a pleomorphic morphology. Although UPS is characterised by complex genetic abnormalities with numerous copy number aberrations, none of these abnormalities are specific to UPS. In this study, three amplified genes, previously identified by our research team as having a potential driver role in UPS tumorigenesis, were functionally analyzed to establish a correlation between the gene expression and the aberrant copy number. Initially, semi-quantitative analysis, by immunocytochemistry and western blotting were performed for YAP1, VGLLl3 and JUN. Based on these analyses, one of these genes, YAP1 was chosen for further study. The effects of YAP1 inhibition on the UPS cell lines, was investigated using a small molecule inhibitor (CA3). The inhibition of YAP1 resulted in a significant reduction of the proliferation of the UPS cells. Moreover, such inhibition resulted in inducing apoptosis in these cells. In the light of these finding, Crispr/Cas9 technology was successfully undertaken on the four UPS cell lines to knock out YAP1. The knockout efficiency was assessed by different approaches, Genomic cleavage detection (GCD) assay, tracking indels by decomposition (TIDE) and Inference of CRISPR Edits (ICE). Consequently, single cell colonies were derived from each cell line to obtain cells with homogeneously edited genomes. For these clones, the impacts of Crispr/Cas9 editing system were evaluated for both mRNA and protein levels. The proliferation, migration and invasion of these clones and their wild-type correspondent were therefore compared to investigate the impact of YAP1 knockout on the development and progression of UPS. Collectively, the findings in this present study suggest a driver role of YAP1 in the UPS tumorigenesis and provide a plausible therapeutic target for UPS treatment.25 0