Characterizing Chromosomal Aberrations in Cells Deficient for Both ATM and MSH2

<p>Ataxia telangiectasia mutated (ATM) and mutS homologue 2 (MSH2) are important DNA repair proteins that participate in DNA repair pathways to maintain genomic integrity. Mice deficient for ATM and MSH2 mice are viable. However, <em>ATM^-/-</em> mice show growth retardation, neurological defects, and spontaneous lymphomagenesis. <em>MSH2^-/-</em> mice suffer from aggressive lymphoid tumors between two to five months of age and have increased microsatellite instability, which predisposes <em>MSH2^-/-</em> mice to carcinomas. However, mice deficient in both ATM and MSH2 are unable to survive beyond postnatal day 21 (P21). The observed lethality in <em>ATM^-/-MSH2^-/-</em> mice may result from the accumulation of genomic instability. To address this hypothesis, metaphase spreads were examined for chromosomal aberrations in B cells, which undergo genetically programmed DNA recombination and mutation. To bypass the lethality in <em>ATM^-/-MSH2^-/- </em>mice, a conditional knockout of ATM was bred onto an <em>MSH2^-/-</em> background strain to generate <em>CD21cre⁺ATM^F/FMSH2^-/-</em> mice, which are referred to as <em>ATM</em><em>^∆/∆</em><em>MSH2^-/-</em>. In these mice, <em>Cre</em> expression, which is regulated by the CD21 promoter, deletes floxed <em>ATM</em> alleles in naïve mature <em>MSH2^-/-</em> B-cells. <em>ATM</em><em>^∆/∆</em> B-cells showed low levels of chromosomal aberrations whereas <em>ATM</em><em>^∆/∆</em><em>MSH2^-/-</em>B-cells had levels of chromosomal aberrations that was comparable to <em>MSH2^-/-</em> B-cells. The latter result suggested that ATM was incompletely deleted in <em>ATM</em><em>^∆/∆</em><em> </em>B-cells. Analysis of <em>ATM</em><em>^∆/∆</em><em> </em>B cell lysates showed residual ATM protein that may account for the comparable chromosomal aberrations in <em>ATM</em><em>^∆/∆</em><em>MSH2^-/-</em>and <em>MSH2^-/-</em>metaphase spreads. <em>ATM^-/-MSH2^-/-</em> embryos die <em>in utero</em> at approximately E16-E20. Thus, mouse embryonic fibroblasts (MEFs) were isolated from E13.5-15.5 <em>ATM^-/-MSH2^-/-</em> embryos and examined for chromosomal aberrations. The <em>ATM^-/-MSH2^-/-</em> MEF metaphase spreads displayed a similar pattern of chromosomal aberrations as <em>ATM^-/-</em> MEFs, which does not support the hypothesis that<em> ATM^-/-MSH2^-/-</em> mice die from increased genomic instability; however, additional metaphase analyses are required to conclusively determine whether genomic instability does or does not impair <em>ATM^-/-MSH2^-/-</em> embryonic development.</p>

Name thesis:degree_name

Master of Science (M.S.)

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Biology

Year dc:date.available

2019

Author dc:creator

• Inalman, Yeliz

Contributors dc:contributor

• Bao Q. Vuong
• Karen Hubbard
• Shireen Saleque

• Class switch recombination
• metaphase spreads
• B-cells
• MEFs
• DNA repair
• genomic instability
• Cancer Biology
• Cell Biology
• Developmental Biology
• Immunopathology
• Molecular Biology
• Molecular Genetics

Repository record dc:identifier

https://academicworks.cuny.edu/cc_etds_theses/778

OAI identifier oai:identifier

oai:academicworks.cuny.edu:cc_etds_theses-1842