Browsing by Subject "DNA damage"

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Involvement of Caspase-2 in Cisplatin-Induced Cell Death in 2008 Ovarian Cancer Cells
(2008-04-01) Adkins, Brett T.; Basu, Alakananda; Berg, Rance E.; Gryczynski, Zygmunt
Adkins, B., Involvement of caspase-2 in cisplatin-induced cell death in 2008 ovarian cancer cells. Master of Science (Molecular Biology and Immunology) April, 2008, 59 pp., 12 illustrations, bibliography, 73 titles. Cisplatin, one of the most effective anticancer drugs in the treatment of ovarian cancer, causes DNA damage and leads to apoptosis. Caspases, a family of cysteine proteases, are essential for the induction of apoptosis. Initiator caspases activate effector caspases to trigger apoptosis. Caspase-2 can function as both an initiator and effector caspase although there are controversies regarding its role in DNA damage-induced apoptosis. Caspase-2 is the only caspase constitutively located in the nucleus, although its function there is unknown. In the present study we have investigated if caspase-2 is important during cisplatin-induced apoptosis and whether cisplatin treatment affects the localization of caspase-2. Caspase-2 depletion suggested that caspase-2 acts upstream of caspase-2 acts upstream of caspase-9 in cisplatin-induced apoptosis. We also made a novel observation that rottlerin, an inhibitor of DNA damage-induced apoptosis, specifically downregulates caspase-2 via the ubiquitin proteamose-mediated pathway. We further show that cisplatin induces caspase-2 translocation out of the nucleus. Moreover, translocation of caspase-2 is more important for cisplatin-induced cell death.
The Regulation of P53 by Protein Kinase C in Anticancer Drug-Induced Apoptosis
(2001-12-01) Johnson, Cassie L.; Basu, Alakananda; Wu, Ming-Chi
Johnson, C., The regulation of p53 by protein kinase C in anticancer drug-induced apoptosis. Master of Science (Microbiology and Immunology), December, 2001. 43 pp., 11 figures, references, 6 titles. The tumor suppression protein p53 has been implicated in DNA damage-induced apoptosis. Previous studies demonstrated that the protein kinase C (PKC) signal transduction pathway regulates apoptosis induced by the DNA damaging agent cisplatin and is deregulated in cisplatin-resistant cells. The present study examined whether PKC influences p53 and, hence, cellular sensitivity to cisplatin. Basal p53 levels were elevated in cisplatin-resistant HeLa (HeLa/CP) cells as compared to parental HeLa cells. Cisplatin further increased p53 levels in HeLA/CP, but not in HeLA cells. However, rottlerin, a PKC-δ inhibitor that prevents cisplatin-induced apoptosis, caused p53 accumulation in HeLa cells treated with cisplatin. Rottlerin stabilized p53 in response to cisplatin in HeLa cells, whereas cisplatin alone was sufficient to stabilize p53 in HeLa/CP cells.
Tolerating DNA Damage: Translesion Polymerase ETA (η) and its regulation in Saccharomyces Cerevisiae
(2008-05-01) Pabla, Ritu; Siede, Wolfram; Alvarez, Rafael; Reeves, Rustin
Pabla, Ritu., Tolerating DNA damage: Translesion polymerase eta (η) and its regulation in Saccharomyces cerevisiae. Doctor of Philosophy (Cell Biology and Genetics), May 2008, 137 pp., 21 illustrations, bibliography, 151 titles. RAD30 gene encoded DNA polymerase eta (Polη) is the only eukaryotic polymerase that can bypass UV-induced thymine-thymine (T-T) dimers in a predominantly error-free manner. The unique ability of reading bulky and geometrically distorted bases in the template makes the polymerase low-fidelity and error-prone for an undamaged template. The purpose of this study is to delineate the mechanism(s) by which activity of Polη is regulated. The increase in RAD30 transcript after UV damage is not reflected at the protein levels. Instead, Polη is monoubiquitinated constitutively. This posttranslational modification is upregulated in G1 phase and downregulated on entry into S phase of the cell-cycle. This downregulation is further accelerated in response to UV induced DNA damage. A missense mutation (L577Q) of the ubiquitin binding domain (UBZ) results in reduced degree of ubiquitination of the mutant protein outside of G1 and a complete failure to stably interact with ubiquitinated substrates. This mutation renders the strain more UV sensitive and mutagenic, a phenotype resembling a complete RAD30 deletion. In other words, UBZ motif and its interaction with ubiquitinated PCNA is critical for Polη function in vivo. In addition to nucleus, the polymerase localizes in mitochondria suggesting its role in damage tolerance in mitochondria. No drastic changes in the localization of polymerase are observed during cell-cycle progression and after UV damage.