Graduate Thesis Or Dissertation
 

Effects of DNA mismatch repair inhibition in Arabidopsis thaliana

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https://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/nz806321t

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  • Genomic instability underlies diseases of unregulated cell growth that result in cancers and developmental abnormalities in humans. Similar genome destabilizing mechanisms are used to create genetic variety in crops for use in breeding and trait development. Errors that occur during DNA replication may cause mutations if they are not corrected before further cell divisions. DNA mismatch repair (MMR) corrects misinsertions and insertion/deletion DNA loop-outs that arise during DNA replication in plants, animals, prokaryotes, and some archaea, all of which incur mutations at rates 100 to 1,000-fold greater when subjected to inherited or somatic-mismatch repair deficiencies. An understanding of the effects of mismatch repair on somatic and germ-line cells in Arabidopsis thaliana is critical to the development of this plant as a model system for the study of genomic instability. Insertions and deletions of multiples of two base pairs in dinucleotide repeat sequences (microsatellites) occur more frequently in the absence of mismatch repair, and the mismatch-repair status of an individual, tissue, or cell may be inferred on the basis of microsatellite mutation frequency. Single-template PCR analysis measured microsatellite mutation frequencies in leaves and shoot-apical-meristem stem cells, and allowed me to address for the first time an important question: Do plants relax mismatch repair in vegetative tissues relative to meristematic germ-line and floral tissue? Analyses of four microsatellite loci in mismatch repair-deficient and wild type plants surprisingly suggest that there is little difference in mismatch repair activity between leaves and seeds. Mismatch-repair-deficient leaves displayed only two-fold higher microsatellite mutation frequency compared to wild type, and wild-type leaves also displayed a two-fold higher microsatellite mutation frequency compared to shoot-apical- meristems. The high frequency of microsatellite mutation in these wildtype tissues is unexpected, and it suggests that plants relax mismatch repair in differentiated tissues while maintaining genetic fidelity in a small set of stem cells in the shoot apical meristem (SAM). Genome sequencing of msh2⁻/⁻ mutation accumulation A. thaliana lines provides an estimated germ-line mutation rate of 3.9 × 10⁻⁷ in the absence of mismatch repair. Comparison of the rates of base substitution mutation per chromosome in mismatch repair-deficient plants with rates reported for wild-type plants suggests mismatch repair is more efficient on chromosome 5 than on chromosomes 1-4. Bias towards G:C → A:T mutations among transitions is maintained but increased nearly 100-fold in the absence of mismatch repair.
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