The function of the ras+ gene of Schizosaccharomyces pombe has been studied by constructing null and activated alleles of this gene. An activated allele (rasVal 12) inhibits conjugation but has no effect on cell growth, entry into stationary phase or sporulation. The phenotype of rasVal 12 is distinct from that caused by elevating the intracellular level of cAMP. This supports the hypothesis that ras of fission yeast does not modulate adenylate cyclase in a manner analogous to S. cerevisiae RAS. Introduction of a human ras sequence into fission yeast cells containing a non‐functional null allele of ras restored the sexual differentiation process thus indicating that the human sequence can complement S. pombe ras. Our data suggest that although ras genes are highly conserved across a considerable evolutionary divide, the cellular function of the ras gene product varies in different organisms.

The ras1‐ mutation of the fission yeast Schizosaccharomyces pombe interferes with sexual differentiation by preventing conjugation and causing inefficient sporulation. From a gene library, we have isolated a gene, byr1+, which when in high copy number restores efficient sporulation to ras1‐ strains. byr1+ encodes a putative 340‐amino acid protein product, the sequence of which strongly suggests that it functions as a protein kinase. Gene disruption experiments show that loss of byr1+ function does not interfere with mitotic growth but it completely prevents both conjugation and sporulation. byr1 is thus another important gene in the sexual differentiation pathway and we believe that at least part of ras1 function is to act directly or indirectly through byr1 to modulate protein phosphorylation.

A new Schizosaccharomyces pombe mutant (rad32) which is sensitive to gamma and UV irradiation is described. Pulsed field gel electrophoresis of DNA from irradiated cells indicates that the rad32 mutant, in comparison to wild type cells, has decreased ability to repair DNA double strand breaks. The mutant also undergoes decreased meiotic recombination and displays reduced stability of minichromosomes. The rad32 gene has been cloned by complementation of the UV sensitive phenotype. The gene, which is not essential for cell viability and is expressed at a moderate level in mitotically dividing cells, has significant homology to the meiotic recombination gene MRE11 of Saccharomyces cerevisiae. Epistasis analysis indicates that rad32 functions in a pathway which includes the rhp51 gene (the S.pombe homologue to S.cerevisiae RAD51) and that cells deleted for the rad32 gene in conjunction with either the rad3 deletion (a G2 checkpoint mutation) or the rad2 deletion (a chromosome stability and potential nucleotide excision repair mutation) are not viable.

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The radiO, radl6, rad2O, and swi9 mutants of the fission yeast Schizosaccharomyces pombe, isolated by their radiation sensitivity or abnormal mating-type switching, have been shown previously to be allelic. We have cloned DNA correcting the UV sensitivity or mating-type switching phenotype of these mutants and shown that the correcting DNA is encompassed in a single open reading frame. The gene, which we will refer to as radi6, is approximately 3 kb in length, contains seven introns, and encodes a protein of 892 amino acids. It is not essential for viability of S. pombe. The predicted protein is the homolog of the Saccharomyces cerevisiae RADI protein, which is involved in an early step in excision-repair of UV damage from DNA. The approximately 30% sequence identity between the predicted proteins from the two yeasts is distributed throughout the protein. Two-hybrid experiments indicate a strong protein-protein interaction between the products of the radl6 and swilO genes of S. pombe, which mirrors that reported for RADJ and RADIO in S. cerevisiae. We have identified the mutations in the four alleles of radl6. They mapped to the N-terminal (radiO), central (rad2O), and C-terminal (radi6 and swi9) regions. The radiO and rad2O mutations are in the splice donor sequences of introns 2 and 4, respectively. The plasmid correcting the UV sensitivity of the rad2O mutation was missing the sequence corresponding to the 335 N-terminal amino acids of the predicted protein. Neither smaller nor larger truncations were, however, able to correct its UV sensitivity.DNA repair is a process of fundamental importance in maintaining the integrity of the genome. In humans, deficiencies in DNA repair result in severe multisystem genetic disorders such as xeroderma pigmentosum and ataxia telangiectasia. In order to counteract the deleterious effects of DNA damage, cells have evolved a series of complex pathways whereby damage of different types can be reversed, removed, or tolerated. Many genes involved in these repair pathways have been cloned from humans and from both budding and fission yeasts. Genes involved in nucleotide excision repair show a striking level of cross-species sequence conservation (18,19). Thus, the cloned human ERCCJ, ERCC2 (XPD), ERCC3 (XPB), ERCC5 (XPG), XPA and XPC genes all share homologies with yeast excision repair genes (3,19,(21)(22)(23)37).Although DNA repair processes have been most extensively studied in the budding yeast Saccharomyces cerevisiae, the fission yeast Schizosaccharomyces pombe is also proving to be an excellent model for higher eukaryotes. Early work on radiation-sensitive mutants of S. pombe suggested the existence of at least two phenotypic groups (29). Mutants in the first group, thought to be involved in excision-repair, are characterized by sensitivity to UV but not y-radiation, caffeine sensitization to UV damage, and pronounced UV-hypermutability. Mutants in the second group are sensitive to both UV and y-radiation and have lost the caffeine sensitization to UV damage. Their mutabil...