Heterogeneous functional Ty1 elements are abundant in the Saccharomyces cerevisiae genome.
Despite the abundance of Ty1 RNA in Saccharomyces cerevisiae, Ty1 retrotransposition is a rare event. To determine whether transpositional dormancy is the result of defective Ty1 elements, functional and defective alleles of the retrotransposon in the yeast genome were quantitated. Genomic Ty1 elements were isolated by gap repair-mediated recombination of pGTy1-H3(delta 475-3944) HIS3, a multicopy plasmid containing a GAL1/Ty1-H3 fusion element lacking most of the gag domain (TYA) and the protease (PR) and integrase (IN) domains. Of 39 independent gap repaired pGTyHIS3 elements isolated, 29 (74%) transposed at high levels following galactose induction. The presence of restriction site polymorphisms within the gap repaired region of the 29 functional pGTyHIS3 elements indicated that they were derived from at least eight different genomic Ty1 elements and one Ty2 element. Of the 10 defective pGTyHIS3 elements, one was a partial gap repair event while the other nine were derived from at least six different genomic Ty1 elements. These results suggest that most genomic Ty1 elements encode functional TYA, PR and IN proteins. To understand how functional Ty1 elements are regulated, we tested the hypothesis that a TYB protein associates preferentially in cis with the RNA template that encodes it, thereby promoting transposition of its own element. A genomic Ty1 mhis3AI element containing either an in-frame insertion in PR or a deletion in TYB transposed at the same rate as a wild-type Ty1mhis3AI allele, indicating that TYB proteins act efficiently in trans. This result suggests in principle that defective genomic Ty1 elements could encode trans-acting repressors of transposition; however, expression of only one of the nine defective pGTy1 isolates had a negative effect on genomic Ty1 mhis3AI element transposition in trans, and this effect was modest. Therefore, the few defective Ty1 elements in the genome are not responsible for transpositional dormancy.
Molecular Genetics Program, Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany 12201-0509.
M J
MJ
Curcio
D J
DJ
Garfinkel
D000595:Amino Acid Sequence; D001483:Base Sequence; D015825:Chromosomes, Fungal; D004254:DNA Nucleotidyltransferases / Q000235:genetics; D004260:DNA Repair; D004251:DNA Transposable Elements; D004271:DNA, Fungal; D010450:Endopeptidases / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics; D005816:Genetic Complementation Test; D016681:Genome, Fungal; D019426:Integrases; D008969:Molecular Sequence Data; D009154:Mutation; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D004251:DNA Transposable Elements; D004271:DNA, Fungal; D005656:Fungal Proteins; D004254:DNA Nucleotidyltransferases; D019426:Integrases; D010450:Endopeptidases