Κατεύθυνση Κλινική Βιοχημεία - Μοριακή Διαγνωστική
Library of the School of Science

2024-06-05

2024

Ougiaridis Konstantinos-Panagiotis

Σίδερης Διαμάντης, Καθηγητής, Τμήμα Βιολογίας, ΕΚΠΑ

«Μελέτη του ρόλου της RNase Κ v14 σε μηχανισμούς απόπτωσης επαγώμενους από οξειδωτικό stress»

Greek

Study of the role of RNase K v14 in oxidative stress- induced apoptosis mechanisms

Oxidative stress has been defined as the imbalance between the production of active oxygen species and the antioxidant defense and contributes significantly to various pathophysiological processes. The cellular response to stress involves a decision between survival or cell death when the damage is severe. Ribonucleases are key enzymes involved in RNA metabolism, one of the crucial steps in controlling gene expression and therefore in controlling cell fate. Various studies have highlighted the role of these enzymes in stressful situations, where they regulate cell responses through the degradation of certain cellular RNA species. Ribonuclease κ belongs to the class of endoribonucleases and at the mRNA level is expressed in all tissues and at all developmental stages. RNASEK is located on the cell surface and endosomes, while its biological role is still unknown, it seems to take part in the operation of the proton pump (V-ATPase), the endocytosis of viruses, the production of piwi-RNAs, also is involved in the process of apoptosis and is a potential cancer biomarker.
The aim of this master thesis is to study the expression levels of the alternative transcript 14 of RNASEK gene in a human cell lines under the influence of oxidizing agents. For this purpose, HEK293T, MCF7 and HTB9 cells were cultured at increasing concentrations of H2O2 and the proteasome inhibitor, Carfilzomib (CFZ) for various incubation periods and the rates of cytotoxicity and apoptosis were determined by the Trypan Blue exclusion technique and the diphenylamine test, respectively. After confirmation of induction of cellular and apoptotic death by oxidative stress, the expression of transcripts was studied by Real Time qPCR reactions, using fluorescent dye SYBR Green I, after RNA isolation and cDNA generation from 160 cell samples. The results were then statistically processed to correlate expression levels with factors. The results of statistical analysis show that expression levels of transcript 14 are increased (8-10 times) at specific concentrations for the 16, 24 and 48 h H2O2 incubation periods. The fact that transcript 14 has low levels of expression under normal conditions, the increase of it under the influence of oxidative stress, which leads the cell to necrotic and / or apoptotic death, indicates its involvement in these processes. In conclusion, it seems that the process of alternative splicing produces transcripts of a gene with different properties, which may or may not be coded and play important roles in cell function, which are worth characterizing.

Science

molecular biology, ribonuclases, RNASE K, oxidative stress

Yes

2

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106

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https://pergamos.lib.uoa.gr/uoa/dl/object/3400504