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RISS 인기검색어
- 검색키워드
- 저자 : Fitz Gerald, Jonathan Nesbit (검색결과 2 건)
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The G1 DNA damage checkpoint in Saccharomyces cerevisiae
Fitz Gerald, Jonathan Nesbit The University of Chicago 2002 해외박사(DDOD)
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Inhibition of cyclin-dependent kinase (CDK) is a conserved downstream effect of DNA damage checkpoint signaling pathways in eukaryotes. This role in the regulation of cell proliferation and prevention of cancer has made CDK a leading target for rational drug design and chemotherapy. It is possible that <italic>S. cerevisiae</italic> CDK, Cdc28, may similarly respond to checkpoint signals, though currently no such Cdc28 function has been described. In this work, 6 point mutations in the <italic>CDC28</italic> gene were isolated that confer DNA damage checkpoint defects. Homology modeling of the <italic>CDC28 </italic> describes a putative N-terminal binding site that may function in this pathway. In these studies it was noticed that 40% of asynchronous wild-type yeast arrest as unbudded cells after 1600 Gy gamma irradiation. This suggests the existence of a novel G1 DNA damage checkpoint. DNA damage in G1 was found to delay onset of replication, bud emergence and spindle pole body duplication and to extend the window of sensitivity to the yeast mating pheromone. These findings all argue that DNA damage can inhibit the Start event in the yeast cell cycle. This G1 arrest is dose-dependent and requires the DNA damage detector <italic> RAD9</italic>, but not the DNA polymerase <italic>POL</italic>ϵ. Whether induced by lethal γ or UV irradiation, the <italic>RAD9</italic>- and <italic> RAD17</italic>-dependent G1 checkpoint arrest can be maintained over 18 hours. Sub-lethal γ irradiation causes haploid yeast to preferentially arrest in G1. This G1 arrest after γ irradiation is prolonged by defects in double-strand break repair, but UV irradiation requires excision repair to initiate an arrest. These results are consistent with single-strand DNA being the initiating signal of checkpoint arrest in G1. The molecular components of the G1 DNA damage checkpoint may provide novel targets for future chemotherapies in the treatment of cancer.
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Diala, Fitz Gerald Iheanyichukwu ProQuest Dissertations & Theses University of Cali 2020 해외박사(DDOD)
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Trichomonas vaginalis, an extracellular, flagellated protozoan parasite, is the etiologic agent for trichomoniasis, the most common non-viral sexually transmitted infection, trichomoniasis. While asymptomatic presentation is commonplace, symptomatic infections typically present as vaginitis and cervicitis in women, and urethritis in men. Only 5-nitroimidazole class of drugs, metronidazole (Mz) and tinidazole, is FDA-approved for treatment of infections. To overcome the knowledge gap in Mz targets in T. vaginalis, we used metronidazole-alkyne analog and we employed copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) “click” reaction to enrich these protein targets. Using tandem mass tag quantitative proteomics, we identified novel protein targets in Mz-sensitive and -resistant parasites. We also determined through activity-based protein profiling that metronidazole binds to cysteine residues and subsequently identified cysteine residues that are bound by metronidazole. As the nature of immune response to T. vaginalis infection appears to vary, we also explored whether T. vaginalis parasites harboring M. hominis, an endosymbiont, induce the production of different cytokines from primary human monocytes compared to parasites that do not harbor the endosymbiont. Indeed, we observe that more cytokines are elaborated in response to M. hominis infected parasites. Together, these studies illuminate our knowledge of this important human pathogen, pharmacologically and immunologically.
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