Understanding The Repair Mechanisms at Ionizing Radiation-induced Damage in The Human Genome

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dc.contributor.advisorSowers, Lawrence C
dc.contributor.committeeMemberMitra, Sankar
dc.contributor.committeeMemberHegde, Muralidhar L
dc.contributor.committeeMemberHazra, Tapas
dc.contributor.committeeMemberPazdrak, Konrad
dc.contributor.committeeMemberWiederhold, Lee R
dc.contributor.committeeMemberPandita, Tej K
dc.creatorDutta, Arijit
dc.creator.orcid0000-0002-5675-0092
dc.date.accessioned2019-03-13T20:24:25Z
dc.date.available2019-03-13T20:24:25Z
dc.date.created2016-05
dc.date.submittedMay 2016
dc.date.updated2019-03-13T20:24:26Z
dc.description.abstractIonizing radiation (IR) such as X-rays induce damage clusters in the genome that include DNA double-strand breaks (DSB) with unligatable dirty ends, along with more frequent oxidized bases and single-strand breaks (SSB). While nonhomologous end joining and homologous recombination are major DSB repair pathways which have been extensively characterized over the past decades, contribution of error-prone alternative end joining (Alt-EJ) at X-ray-induced DNA damage is poorly characterized and underestimated. Moreover, how repair of oxidative base lesions and DSB are coordinated at damage clusters is an important unanswered question. I used recircularization of linearized plasmid reporters to monitor repair of DSBs with 3P-blocked termini, which mimic X-ray-induced strand breaks, both in cell and in vitro, with repair complexes and measured relative efficiency of NHEJ vs. Alt-EJ based on sequence analysis of the joint site. Although NHEJ was the predominant pathway for DSB repair, Alt-EJ was significantly enhanced in pre-irradiated cells. This stimulation was dependent on XRCC1 phosphorylation by casein kinase 2 (CK2) that enhanced the interaction of XRCC1 with the end resection enzymes Mre11 and CtIP. The XRCC1 immunocomplex isolated from U2OS cells had Alt-EJ activity in vitro; this activity was significantly higher in the immunocomplex from pre-irradiated cells. Our studies thus suggest that activation of Alt-EJ proficient repair complexes after irradiation in surviving cells could contribute to radioresistance and could be therapeutically targeted. In a separate study, we showed that there is a hierarchy in repair of DSBs by NHEJ followed by base excision repair of oxidized bases at IR-induced damage clusters, coordinated by scaffold attachment factor-A (SAF-A), that is crucial to maintain genomic integrity.
dc.format.mimetypeapplication/pdf
dc.identifier.urihttps://hdl.handle.net/2152.3/11146
dc.subjectClustered DNA damage
dc.subjectIonizing radiation
dc.subjectDouble strand break
dc.subjectNon-homologous end joining
dc.subjectAlternative end joining
dc.subjectGenomic instability
dc.subjectRepair complex
dc.titleUnderstanding The Repair Mechanisms at Ionizing Radiation-induced Damage in The Human Genome
dc.typeThesis
dc.type.materialtext
thesis.degree.departmentBiochemistry and Molecular Biology
thesis.degree.grantorThe University of Texas Medical Branch at Galveston
thesis.degree.levelDoctoral
thesis.degree.nameBiochemistry and Molecular Biology (Doctoral)

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