A molecular and genetic analysis of variable number tandem repeat (VNTR) recombination in Saccharomyces cerevisiae

Eric James Weaver, Thomas Jefferson University

Abstract

One type of repeated sequence present in eukaryotic genomes is the variable number tandem repeat (VNTR). These highly polymorphic sequences consist of reiterated arrays of monomer units that can vary in length and base composition. Because they are so variable, VNTR loci are important to genetic and forensic studies, and some have been implicated in genomic rearrangements associated with disease. To better understand the mechanisms and determinants of VNTR instability, we have developed a Saccharomyces cerevisiae system to examine mitotic direct repeat deletion-recombination involving a human VNTR sequence. We found that this sequence was unstable in S. cerevisiae and we noted that identical VNTR arrays recombined at higher rates than those divergent in sequence by 17%. We also noted that this spontaneous recombination was dependent upon the recombination/DNA repair gene RAD52. Other recombination/DNA repair genes RAD1 and PMS1 appear to have relatively little involvement in VNTR deletion. We observed that VNTR instability is increased up to 10-fold in strains that are deficient in DNA topoisomerase III function. Other known suppressors of recombination, TOP1 and HPR1 did not appear to influence the stability of VNTR sequences in our system. We found that deletions involving diverged (homeologous) VNTR sequences generated a distribution of different sized recombination products potentially resulting from discrete alignments of VNTR array monomers during recombination. The frequency of recovery of individual products correlated with the amount of sequence identity characterizing the associated VNTR alignments. Deletions involving identical VNTR arrays resulted primarily in products of a single size, implying usage of a single preferred alignment of these sequences (the maximum alignment) during recombination even though other less homologous alignments are possible. Double-strand break-induced VNTR recombination resulted in a similar profile of recombination products. However, this type of event was equally frequent in identical and diverged sequences, and was less dependent than analogous spontaneous events on RAD52 function. These results suggest that the imperfect internal redundancy of a VNTR sequence produces a milieu of competing recombination modes characterized by differing amounts of sequence homology; the amount of homology associated with a given mode appears to influence the frequency of its usage. The RAD52-dependence of spontaneous mitotic VNTR sequence deletion in our system is consistent with proposed models of human VNTR recombination that invoke gene conversion. The ability of VNTRs to rearrange via a RAD52-independent pathway in our double-strand break-induced recombination assay also suggests a role for RAD52-independent recombination mechanisms such as single-strand annealing in the instability of these complex sequences.

Subject Area

Genetics|Molecular biology|Microbiology

Recommended Citation

Weaver, Eric James, "A molecular and genetic analysis of variable number tandem repeat (VNTR) recombination in Saccharomyces cerevisiae" (1996). ETD Collection for Thomas Jefferson University. AAI9633534.
https://jdc.jefferson.edu/dissertations/AAI9633534

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