Document Details
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Document Type |
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Thesis |
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Document Title |
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Molecular Profiling of Epitranscriptomes in Extracellular Matrix Detached Cancer Cells التنميط الجزيئي للإيبيترانسكربتوم في النسيج البيني خارج الخلايا السرطانية المنفصلة |
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Subject |
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Faculty of Science |
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Document Language |
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Arabic |
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Abstract |
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Introduction & Aim: Epitranscriptomes are modifications on RNA transcripts that promote RNA metabolisms such as RNA stability, RNA-mediated decay, RNA splicing, and translational efficiency. These epitranscriptomes are added either by writers (methyl- and acetyl-transferases), removed by erasers (demethylase and de-acetylase) or bound by readers (RNA binding proteins). Collectively, the writers, erasers, and readers are referred to as epitranscriptomic regulators. Aberrant levels of epitranscriptomes and their regulators have been reported in diseases such as diabetes, bacterial infection, viral infection, and cancer. In cancer, epitranscriptomes are associated with cell proliferation, cell growth, epithelial-to-mesenchymal transition (EMT), metastasis, and stemness.
Extracellular matrix (ECM) detached cancer cells were recently reported to possess metastatic and stemness properties, therefore, in this study epitranscriptomic regulators were profiled in ECM detached cancer cells, and amongst all the profiled epitanscriptomic regulators; N-acetyltransferase 10 (NAT10) was found to be consistently overexpressed in all ECM detached cancer cells. NAT10 is an RNA cytidine transferase that adds an acetyl group at the 4th position of cytidine in RNA transcript including mRNA, tRNA, and rRNA resulting in RNA stability, rRNA biogenesis, and translational efficiency. In cancer, NAT10 promotes cell proliferation, cell growth, EMT, and poor patient survival. This study utilized multi-omics approaches such as untargeted metabolomics, lipidomics, transcriptomics as well as epitranscriptomics to identify NAT10-mediated key molecules and profile pathways that could be targeted for diagnostic and therapeutic purposes.
Methods: High-throughput RNA sequencing, acetylated-RNA immunoprecipitation-sequencing (acRIP-seq) and untargeted metabolomics and lipidomics were performed in NAT10 depleted cancer cells. Further, functional analysis using flow cytometer was conducted for apoptosis, cell cycle, mitochondrial membrane potential, reactive oxygen species (ROS), and lipid ROS assays.
Results: Through lipidomic and metabolomic analysis, NAT10 and its inhibitor; Remodelin were found to modulate fatty acid (FA) metabolism and mitochondrial lipid metabolism respectively. Additionally, ferroptosis was observed to be induced in NAT10-depleted cancer cells as well as Remodelin treated cancer cells. Transcriptomic data showed downregulation of lipid metabolism and ferroptosis regulatory genes such as ELOVL6, ACSL1, ACSL3, ACSL4, ACADSB, ACAT1, GCLC, SLC7A11, MAP1LC3A, and SLC39A8 in NAT10 depleted and Remodelin treated cancer cells. Consistently, epitranscriptomic data showed a reduction in ac4C levels on the mRNA transcripts of the FA-related genes (ELOVL6, ACSL1, ACSL3, and ACSL4) and ferroptosis-related genes (GCLC, SLC7A11). Flow cytometry and biochemical analysis further confirmed evidence on the impact of NAT10 on cancer survival and lipid modulation through measurement of cell death, cell cycle, mitochondrial potential, ROS, antioxidant activities, lipid ROS, and lipid droplet formation. Collectively showing high ROS, lipid ROS and depolarized mitochondria in NAT10 depleted and Remodelin treated cancer cells implicated in elevated cell death via apoptosis and ferroptosis and cell cycle arrest.
Conclusion: Conclusively, the results provide novel insights into the critical role of NAT10 as a regulatory factor in lipid metabolic reprogramming and showed the benefit of targeting NAT10 for cancer treatment. |
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Supervisor |
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Prof. Hani Choudhry |
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Thesis Type |
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Doctorate Thesis |
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Publishing Year |
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1444 AH
2023 AD |
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Co-Supervisor |
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Dr. Mohammad Imran Khan |
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Added Date |
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Tuesday, July 18, 2023 |
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Researchers
| محمود حسن دلهات | Dalhat, Mahmood Hassan | Researcher | Doctorate | |
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