Abstract:
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.
