Homologous recombination hotspots increase the frequency of recombination in nearby DNA. The M26 hotspot in the ade6 gene of Schizosaccharomyces pombe is a meiotic hotspot with a discrete, cis-acting nucleotide sequence (5-ATGACGT-3) defined by extensive mutagenesis. A heterodimeric M26 DNA binding protein, composed of subunits Mts1 and Mts2, has been identified and purified 40,000-fold. Cloning, disruption, and genetic analyses of the mts genes demonstrate that the Mts1͞Mts2 heterodimer is essential for hotspot activity. This provides direct evidence that a specific trans-acting factor, binding to a cis-acting site with a unique nucleotide sequence, is required to activate this meiotic hotspot. Intriguingly, the Mts1͞Mts2 protein subunits are identical to the recently described transcription factors Atf1 (Gad7) and Pcr1, which are required for a variety of stress responses. However, we report differential dependence on the Mts proteins for hotspot activation and stress response, suggesting that these proteins are multifunctional and have distinct activities. Furthermore, ade6 mRNA levels are equivalent in hotspot and nonhotspot meioses and do not change in mts mutants, indicating that hotspot activation is not a consequence of elevated transcription levels. These findings suggest an intimate but separable link between the regulation of transcription and meiotic recombination. Other studies have recently shown that the Mts1͞Mts2 protein and M26 sites are involved in meiotic recombination elsewhere in the S. pombe genome, suggesting that these factors help regulate the timing and distribution of homologous recombination.

Bähler et al.(1998) recently described a PCR‐based system for the deletion, tagging and overexpression of endogenous genes in the fission yeast Schizosaccharomyces pombe. A small set of PCR primers can be used to generate gene‐targeting substrates from each of several modules that differ in the selectable marker (ura4+ or kanMX6), the presence or absence of specific epitope tags (HA, Myc, GST or GFP), the position in which the epitopes will be inserted (C‐ or N‐terminal), and the presence or absence of a regulatable promoter (the nmt1 promoter). This is a straightforward and powerful system: nine different genes were C‐terminal tagged at an average efficiency of 73%, using primers producing only 60–81 bp of homology. In contrast, when studying three transcriptionally‐silent genes (rec8+, rec10+ and rec11+) we obtained an average homologous integration efficiency of 4% for 12 targeting constructs when using primers that contained 80 bp of homology. By using a PCR‐based increase in the amount of flanking homology to ≥250 bp, we obtained homologous integration efficiencies of up to 100%. Thus, loci of S. pombe that are refractory to gene targeting when using short tracts of homology can be readily modified by increasing the extent of homology flanking the targeting modules. This straightforward and cost‐effective approach might therefore be the one of choice for the modification of S. pombe loci in general and of targeting‐refractory loci in particular. Copyright © 1999 John Wiley & Sons, Ltd.

Homologous recombination hot spots are cis-acting DNA sites that increase the frequency of recombination in their vicinity and thereby influence patterns of genomic recombination (22, 44). Mechanisms of recombination initiation involving double-strand DNA breaks (3,5,18,28,40), regulation of recombination involving competition between nearby hot spots

During meiosis homologous chromosomes replicate once, pair, experience recombination, and undergo two rounds of segregation to produce haploid meiotic products. The rec8+, rec10+, and rec11+ genes of the fission yeast Schizosaccharomyces pombe exhibit similar specificities for meiotic recombination and rec8+ is required for sister chromatid cohesion and homolog pairing. We applied cytological and genetic approaches to identify potential genetic interactions and to gauge the fidelity of meiotic chromosome segregation in the mutants. The rec8+ gene was epistatic to rec10+ and to rec11+, but there was no clear epistatic relationship between rec10+ and rec11+. Reciprocal (crossover) recombination in the central regions of all three chromosomes was compromised in the rec mutants, but recombination near the telomeres was nearly normal. Each of the mutants also exhibited a high rate of aberrant segregation for all three chromosomes. The rec8 mutations affected mainly meiosis I segregation. Remarkably, the rec10 and rec11 mutations, which compromised recombination during meiosis I, affected mainly meiosis II segregation. We propose that these genes encode regulators or components of a “meiotic chromatid cohesion” pathway involved in establishing, maintaining, and appropriately releasing meiotic interactions between chromosomes. A model of synergistic interactions between sister chromatid cohesion and crossover position suggests how crossovers and cohesion help ensure the proper segregation of chromosomes in each of the meiotic divisions.