The Strong ADH1 Promoter Stimulates Mitotic and Meiotic Recombination at the ADE6 Gene of Schizosaccharomyces pombe

Abstract

The effect of the strong promoter from the alcohol dehydrogenase gene on mitotic and meiotic intragenic recombination has been studied at the ade6 locus of the fission yeast Schizosaccharomyces pombe. A 700-bp fragment containing the functional adh1 promoter was used to replace the weak wild-type promoter of the ade6 gene. Analysis of mRNA showed that strains with this ade6::adh1 fusion construct had strongly elevated ade6-specific mRNA levels during vegetative growth as well as in meiosis. These increased levels of mRNA correlated with a 20- to 25-fold stimulation of intragenic recombination in meiosis and a 7-fold increased prototroph formation during vegetative growth. Analysis of flanking marker configurations of prototrophic recombinants indicated that simple conversions as well as conversions associated with crossing over were stimulated in meiosis. The strongest stimulation of recombination was observed when the adh1 promoter was homozygous. Studies with heterologous promoter configurations revealed that the highly transcribed allele was the preferred acceptor of genetic information. The effect of the recombinational hot spot mutation ade6-M26 was also investigated in this system. Its effect was only partly additive to the elevated recombination rate generated by the ade6::adh1 fusion construct.

A protein complex containing Tho2, Hpr1, Mft1 and a novel protein, Thp2, connects transcription elongation with mitotic recombination in Saccharomyces cerevisiae

“…In yeast, the ®nding that HOT1, a cisacting hotspot of recombination, was dependent on RNA polymerase I (RNAPI)-driven transcription (Voelkel-Meiman et al, 1987;Stewart and Roeder, 1989) provided the ®rst example of transcription-induced recombination. Subsequently, similar observations were made for RNA polymerase II (RNAPII)-driven transcription (Thomas and Rothstein, 1989;Grimm et al, 1991;Nevo-Caspi and Kupiec, 1994). In mammalian cells, RNAPII-driven transcription has been shown to stimulate homologous recombination (Nickoloff, 1992;Thyagarajan et al, 1995) and to positively control V(D)J recombination (Blackwell et al, 1986;Lauster et al, 1993;Oltz et al, 1993) and class switching (Daniels and Lieber, 1995).…”

Transcription-Associated Genome Instability

“…In agreement with the hypothesis that the level of transcription does matter for TAR, replacement of the weak ADE6 promoter by the strong ADH1 promoter in S. pombe was shown to cause a 7-fold increase in intragenic recombination between two ade6 alleles. 127 One decade later, the impact of RNAPII transcription on different kinds of recombination processes, such as simple gene conversion or crossover-associated events, was analyzed using interchromosomal systems based on different lys2 alleles located on nonhomologous chromosomes and controlled by promoters of different strength. 128 High transcription of both lys2 alleles led to a 3.8-fold increase in recombination frequency as compared to low transcription, with the majority of recombinants having undergone a reciprocal crossover event.…”

The connection between transcription and genomic instability

“…In yeast, RNA polymerase II (RNAPII)‐mediated TAR was first shown in direct repeats transcribed under the control of the regulatable GAL1 promoter Induction of transcription increased deletions by ∼10‐fold (Thomas and Rothstein, 1989) TAR has also been reported in other recombination assays in S.cerevisiae (Nevo‐Caspi and Kupiec, 1994; Bratty et al ., 1996; Saxe et al ., 2000), Schizosaccharomyces pombe (Grimm et al ., 1991) and mammalian cells (Nickoloff, 1992) or in V(D)J recombination (Blackwell et al ., 1986; Lauster et al ., 1993) Therefore, transcription may produce structures that are not only mutagenic but also recombinogenic in all organisms from bacteria to humans.…”