Inhibition of Ty1 transposition by mating pheromones in Saccharomyces cerevisiae.

Abstract

The Tyl elements in the yeast Saccharomyces cerevisiae are a family of retrotransposons which transpose via a process similar to that of retroviral replication. We report here that the Tyl transposition process can be blocked posttranscriptionally by treatment of cells with mating pheromones. When haploid yeast cells are treated with appropriate mating pheromones, the transposition frequency of a marked Tyl element driven by the GAL] promoter is greatly diminished. Tyl viruslike particles (VLPs), the putative intermediates for transposition, can be isolated from mating pheromone-treated cells. These VLPs accumulate to normal levels but are aberrant in that they produce very few reverse transcripts of Tyl RNA both in vivo and in vitro and contain subnormal amounts of p90-TYB and related proteins. In addition, a TYA phosphoprotein product accumulates in treated cells, and some species of TYB proteins have decreased stability. We also show that decreased transposition in mating pheromone-treated cells is not a consequence of simply blocking cell division, since Tyl transposes at a nearly normal rate in yeast cells arrested in G2 by the drug nocodazole.Transposable elements carry out the breakage of host DNA and the insertion of new DNA. Clearly, the frequency of these potentially deleterious reactions must be stringently regulated. In Escherichia coli, the well-studied transposon TnJO is regulated transcriptionally, translationally, and at the level of the transposition reaction itself (21). In yeast cells, the transposition of Tyl elements is known to be controlled transcriptionally both by the MAT locus and by several SPT loci, yet examples of posttranscriptional regulatory mechanisms are few (3).Tyl elements, a family of retrotransposons in the yeast Saccharomyces cerevisiae, are approximately 6 kb long and consist of a central coding region flanked by long terminal repeats. The two open reading frames in Tyl include TYA, which corresponds to retroviral gag, and TYB, which corresponds to pol (3). Previous studies indicate that Tyl elements transpose via a mechanism analogous to that of retroviral replication and that the functions and relationships of Tyl-encoded proteins are also very similar to those of retroviral proteins (1,5,26,36). Specifically, TYB proteins are initially synthesized as a fusion product of the TYA and TYB open reading frames (p190-TYA/TYB). Both TYB and TYA precursor proteins are processed by a Tyl-encoded protease to smaller products (Fig. 1); this processing is essential for transposition (36). It has been shown that TYA proteins are phosphorylated, although it is not known whether the phosphorylation plays any role in transposition (23). Viruslike particles (VLPs) have been isolated from yeast cells in which Tyl RNAs are overexpressed (19,24). VLPs are apparently direct intermediates for transposition; they are capable of reverse transcribing Tyl RNA and integrating Tyl cDNA, two key steps in the life cycle of Tyl elements (15,16,19,24).

Yeast Ty1 Retrotransposition Is Stimulated by a Synergistic Interaction between Mutations in Chromatin Assembly Factor I and Histone Regulatory Proteins

“…We could not determine if the mutation was recessive for the high transposition rate phenotype because Ty1 transcription is down-regulated in MATa/ MAT␣ diploid cells (52,75).…”

“…Processing and immunogold labeling were performed as described elsewhere (2) with the changes noted below. Cells fixed overnight at 4°C in buffer (4% sucrose in 100 mM sodium phosphate [pH 7.5]) containing 1% acrolein and 4% paraformaldehyde were washed twice with this same buffer for 10 min and then dehydrated at room temperature in an ethylene glycol concentration gradient (25,50,75,90, and 90% for 15, 15, 15, 10, and 10 min, respectively). Infiltration was carried out over a period of 3 days.…”

Ty1 Proteolytic Cleavage Sites Are Required for Transposition: All Sites Are Not Created Equal

“…The transposition assays were done using congenic yeast strains YH10 (MATa ura3-52 his4-539 lys2-801 GAL ϩ ) (38) and YH51 (MATa ura3-52 his4-539 lys2-801 spt3 GAL ϩ ). Cell extracts were prepared from YH51 strains carrying plasmids with mutant Gal-Ty1-neo constructs.…”

Ty3, a Position-specific Retrotransposon in Budding Yeast

“…Despite common upstream components of these pathways, cross talk between them is controlled by mating MAP kinase Fus3-mediated degradation of Tec1 in haploid cells and mating type suppression of Ty3 in diploid cells (33). Ty1 constitutes a high fraction of RNA in haploid cells (34), but Ty1 proteins are actively degraded during the mating pheromone response (35)(36)(37). Hence, despite overlapping induction pathways and shared host factors, Ty1 and Ty3 occupy discrete retrotransposition niches.…”