A Regulatory Role for ATM in Suppression of Mre11-Dependent DNA Degradation and Microhomology-Mediated End Joining

Persistent Link:
http://hdl.handle.net/10150/194399
Title:
A Regulatory Role for ATM in Suppression of Mre11-Dependent DNA Degradation and Microhomology-Mediated End Joining
Author:
Rahal, Elias Adel
Issue Date:
2009
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
ATM is the defective kinase in the neurodegenerative disorder ataxia telangiectasia. This kinase is associated with DNA double-strand break (DSB) repair and cell cycle control. Our laboratory previously demonstrated elevated levels of deletions and error-prone double-strand break repair via microhomology-mediated end joining (MMEJ) in ATM-deficient (A-T) extracts when compared to controls (wtATM+). To assess the involvement of enhanced nuclease activities in A-T extracts we studied the stability of DNA duplex substrates in A-T and control nuclear extracts under DSB repair conditions. We observed a marked shift in detection from full-length products to shorter products in A-T extracts. Addition of purified ATM to A-T nuclear extracts restored full-length product detection. This repression of degradation by ATM was dependent on its kinase activities. These results demonstrated a role for ATM in suppressing the degradation of DNA ends possibly through inhibiting nucleases implicated in MMEJ such as Mre11. Therefore, we assessed DNA end-stability in Mre11-depleted nuclear extracts and in extracts treated with the Mre11 nuclease inhibitor, Mirin. This resulted in decreased DNA degradation in both control and A-T extracts. Knockdown of Mre11 levels also led to an enhancement of DNA end-stability in nuclear extracts. Examining MMEJ levels by employing an in vivo reporter assay system revealed a decline in this pathway in Mre11-knockdown cells and in those treated with Mirin. These results signify a role for the Mre11 nuclease in MMEJ in mammalian cells and indicate a regulatory function for ATM in the control of error-prone DSB repair and preservation of DNA end-stability at a break.
Type:
text; Electronic Dissertation
Keywords:
Ataxia Telangiectasia; ATM; DNA repair; Microhomology-Mediated End Joining; Mre11; MRN
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Molecular & Cellular Biology; Graduate College
Degree Grantor:
University of Arizona
Advisor:
Dixon, Kathleen

Full metadata record

DC FieldValue Language
dc.language.isoENen_US
dc.titleA Regulatory Role for ATM in Suppression of Mre11-Dependent DNA Degradation and Microhomology-Mediated End Joiningen_US
dc.creatorRahal, Elias Adelen_US
dc.contributor.authorRahal, Elias Adelen_US
dc.date.issued2009en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractATM is the defective kinase in the neurodegenerative disorder ataxia telangiectasia. This kinase is associated with DNA double-strand break (DSB) repair and cell cycle control. Our laboratory previously demonstrated elevated levels of deletions and error-prone double-strand break repair via microhomology-mediated end joining (MMEJ) in ATM-deficient (A-T) extracts when compared to controls (wtATM+). To assess the involvement of enhanced nuclease activities in A-T extracts we studied the stability of DNA duplex substrates in A-T and control nuclear extracts under DSB repair conditions. We observed a marked shift in detection from full-length products to shorter products in A-T extracts. Addition of purified ATM to A-T nuclear extracts restored full-length product detection. This repression of degradation by ATM was dependent on its kinase activities. These results demonstrated a role for ATM in suppressing the degradation of DNA ends possibly through inhibiting nucleases implicated in MMEJ such as Mre11. Therefore, we assessed DNA end-stability in Mre11-depleted nuclear extracts and in extracts treated with the Mre11 nuclease inhibitor, Mirin. This resulted in decreased DNA degradation in both control and A-T extracts. Knockdown of Mre11 levels also led to an enhancement of DNA end-stability in nuclear extracts. Examining MMEJ levels by employing an in vivo reporter assay system revealed a decline in this pathway in Mre11-knockdown cells and in those treated with Mirin. These results signify a role for the Mre11 nuclease in MMEJ in mammalian cells and indicate a regulatory function for ATM in the control of error-prone DSB repair and preservation of DNA end-stability at a break.en_US
dc.typetexten_US
dc.typeElectronic Dissertationen_US
dc.subjectAtaxia Telangiectasiaen_US
dc.subjectATMen_US
dc.subjectDNA repairen_US
dc.subjectMicrohomology-Mediated End Joiningen_US
dc.subjectMre11en_US
dc.subjectMRNen_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineMolecular & Cellular Biologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorDixon, Kathleenen_US
dc.contributor.committeememberWeinert, Teden_US
dc.contributor.committeememberBosco, Giovannien_US
dc.contributor.committeememberSchroeder, Joyceen_US
dc.contributor.committeememberZarnescu, Danielaen_US
dc.identifier.proquest10286en_US
dc.identifier.oclc659750907en_US
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