Elisabeth Lehmann

Rec8p, a meiotic recombination and sister chromatid cohesion phosphoprotein of the Rad21p family conserved from fission yeast to humans.

ABSTRACT Our work and that of others defined mitosis-specific (Rad21 subfamily) and meiosis-specific (Rec8 subfamily) proteins involved in sister chromatid cohesion in several eukaryotes, including humans. Mutation of the fission yeast Schizosaccharomyces pombe rec8 gene was previously shown to confer a number of meiotic phenotypes, including strong reduction of recombination frequencies in the central region of chromosome III, absence of linear element polymerization, reduced pairing of homologous chromosomes, reduced sister chromatid cohesion, aberrant chromosome segregation, defects in spore formation, and reduced spore viability. Here we extend the description of recombination reduction to the central regions of chromosomes I and II. We show at the protein level that expression ofrec8 is meiosis specific and that Rec8p localizes to approximately 100 foci per prophase nucleus. Rec8p was present in an unphosphorylated form early in meiotic prophase but was phosphorylated prior to meiosis I, as demonstrated by analysis of the mei4mutant blocked before meiosis I. Evidence for the persistence of Rec8p beyond meiosis I was obtained by analysis of the mutantmes1 blocked before meiosis II. A human gene, which we designate hrec8, showed significant primary sequence similarity to rec8 and was mapped to chromosome 14. High mRNA expression of mouse and human rec8 genes was found only in germ line cells, specifically in testes and, interestingly, in spermatids. hrec8 was also expressed at a low level in the thymus. Sequence similarity and testis-specific expression indicate evolutionarily conserved functions of Rec8p in meiosis. Possible roles of Rec8p in the integration of different meiotic events are discussed.

Novel Genes Required for Meiotic Chromosome Segregation Are Identified by a High-Throughput Knockout Screen in Fission Yeast

Two rounds of chromosome segregation after only a single round of DNA replication enable the production of haploid gametes from diploid precursors during meiosis. To identify genes involved in meiotic chromosome segregation, we developed an efficient strategy to knock out genes in the fission yeast on a large scale. We used this technique to delete 180 functionally uncharacterized genes whose expression is upregulated during meiosis. Deletion of two genes, sgo1 and mde2, caused massive chromosome missegregation. sgo1 is required for retention of centromeric sister-chromatid cohesion after anaphase I. We show here that mde2 is required for formation of the double-strand breaks necessary for meiotic recombination.

Involvement of nucleotide-excision repair in msh2 pms1 -independent mismatch repair

Nucleotide-excision repair (NER) and mismatch repair (MMR) are prominent examples of highly conserved DNA repair systems which recognize and replace damaged and/or mispaired nucleotides in DNA. In humans, inheritable defects in components of the NER system are associated with severe diseases such as xeroderma pigmentosum (XP) and Cockayne syndrome (CS), whereas inactivation of MMR is accompanied by predisposition to certain types of cancer. In Schizosaccharomyces pombe, the msh2- and pms1-dependent long-patch MMR system efficiently corrects small insertion/deletion loops and all base-base mismatches, except C/C. Up to 70% of C/C mismatches generated in recombination intermediates, and to a lesser extent also other base-base mismatches, are thought to undergo correction by a minor, short-patch excision repair system. We identify here the NER genes rhp14, swi10 and rad16 as components of this repair pathway and show that they act independently of msh2 and pms1.

The msh2 Gene of Schizosaccharomyces pombe Is Involved in Mismatch Repair, Mating-Type Switching, and Meiotic Chromosome Organization

ABSTRACT We have identified in the fission yeast Schizosaccharomyces pombe a MutS homolog that shows highest homology to the Msh2 subgroup. msh2 disruption gives rise to increased mitotic mutation rates and increased levels of postmeiotic segregation of genetic markers. In bandshift assays performed with msh2Δ cell extracts, a general mismatch-binding activity is absent. By complementation assays, we showed that S. pombe msh2 is allelic with the previously identified swi8 andmut3 genes, which are involved in mating-type switching. The swi8-137 mutant has a mutation in the msh2gene which causes a truncated Msh2 peptide lacking a putative DNA-binding domain. Cytological analysis revealed that during meiotic prophase of msh2-defective cells, chromosomal structures were frequently formed; such structures are rarely found in the wild type. Our data show that besides having a function in mismatch repair,S. pombe msh2 is required for correct termination of copy synthesis during mating-type switching as well as for proper organization of chromosomes during meiosis.

Roles of Hop1 and Mek1 in Meiotic Chromosome Pairing and Recombination Partner Choice in Schizosaccharomyces pombe

ABSTRACT Synaptonemal complex (SC) proteins Hop1 and Mek1 have been proposed to promote homologous recombination in meiosis of Saccharomyces cerevisiae by establishment of a barrier against sister chromatid recombination. Therefore, it is interesting to know whether the homologous proteins play a similar role in Schizosaccharomyces pombe. Unequal sister chromatid recombination (USCR) was found to be increased in hop1 and mek1 single and double deletion mutants in assays for intrachromosomal recombination (ICR). Meiotic intergenic (crossover) and intragenic (conversion) recombination between homologous chromosomes was reduced. Double-strand break (DSB) levels were also lowered. Notably, deletion of hop1 restored DSB repair in rad50S meiosis. This may indicate altered DSB repair kinetics in hop1 and mek1 deletion strains. A hypothesis is advanced proposing transient inhibition of DSB processing by Hop1 and Mek1 and thus providing more time for repair by interaction with the homologous chromosome. Loss of Hop1 and Mek1 would then result in faster repair and more interaction with the sister chromatid. Thus, in S. pombe meiosis, where an excess of sister Holliday junction over homologous Holliday junction formation has been demonstrated, Hop1 and Mek1 possibly enhance homolog interactions to ensure wild-type level of crossover formation rather than inhibiting sister chromatid interactions.

Genetic analysis of particular aspects of intergenic conversion in Schizosaccharomyces pombe

SummaryThe phenomenon of intergenic conversion (information transfer between dispersed but sequence-related genetic elements embedded in non homologous environments) has been demonstrated clearly in S. pombe as well as in other organisms. By analysing four particular aspects of intergenic conversion in S. pombe we reach the following conclusions: (1) The events are not RNA-mediated since transcription-deficient members of the serine tRNA gene family studied are as productive in intergenic information transfer as transcription-proficient members, (2) Intergenic recombination is not a reciprocal but a non-reciprocal process, (3) Intergenic conversion occurring at a particular locus does not influence crossing-over in the intervals adjacent to that locus, and (4) During meiosis the process occurs at the replicated (chromatid) stage rather than at the unreplicated (chromosome) stage.

Concerning the map of chromosome I of Schizosaccharomyces pombe

SummaryThe correct orientation of a large segment (at least 200 cM) on the left arm of chromosome I of S. pombe is inverted relative to the one given in a recent mapping paper.