Characterization offus1ofSchizosaccharomyces pombe: a Developmentally Controlled Function Needed for Conjugation

Abstract

In Schizosaccharomyces pombe, the fus1 mutation blocks conjugation at a point after cell contact and agglutination. The cell walls separating the mating partners are not degraded, which prevents cytoplasmic fusion. In order to investigate the molecular mechanism of conjugation, we cloned the fus1 gene and found that it is capable of encoding a 1,372-amino-acid protein with no significant similarities to other known proteins. Expression of the fus1 gene is regulated by the developmental state of the cells. Transcription is induced by nitrogen starvation and requires a pheromone signal in both P and M cell types. Consequently, mutants defective in the pheromone response pathway fail to induce fus1 expression. The ste11 gene, which encodes a transcription factor controlling expression of many genes involved in sexual differentiation, is also required for transcription of fus1. Furthermore, deletion of two potential Ste11 recognition sites in the fus1 promoter region abolished transcription, and expression could be restored when we inserted a different Ste11 site from the mat1-P promoter. Since this element was inverted relative to the fus1 element, we conclude that activation of transcription by Ste11 is independent of orientation. Although the fus1 mutant has a phenotype very similar to that of Saccharomyces cerevisiae fus1 mutants, the two proteins appear to have different roles in the process of cell fusion. Budding yeast Fus1 is a typical membrane protein and contains an SH3 domain. Fission yeast Fus1 has no features of a membrane protein, yet it appears to localize to the projection tip. A characteristic proline-rich potential SH3 binding site may mediate interaction with other proteins.Nutritional starvation is the major signal that activates sexual differentiation in the fission yeast Schizosaccharomyces pombe (11). As long as the nutritional conditions are favorable, haploid cells will propagate vegetatively, but under conditions of nitrogen starvation, the cells exit from the mitotic cycle and undergo a differentiation process, which requires sexual agglutination, conjugation, nuclear fusion, meiosis, and spore formation to occur in an orderly fashion (see reference 13). The process of conjugation involves the action of diffusible pheromones secreted by P and M cell types in order to attract each other. When exposed to the opposite pheromone, the cells form projections toward each other (18, 37) and fuse upon cell-cell contact. Attachment at the projection tips between paired cells culminates in localized cell wall degradation and plasma membrane fusion. Nuclear fusion is coordinated with these events, resulting in the formation of a zygote (see reference 53).In the differentiation process, the pheromones act by binding to specific receptors on the surface of the opposite cell type (29, 60), thereby activating the pheromone response pathway. Transmission of the signal through the pathway involves the actions of the ras1 function and of three protein kinases encoded by byr2, byr1, and spk1 (19,45,47,49,58,6...

Membrane Organization and Cell Fusion During Mating in Fission Yeast Requires Multipass Membrane Protein Prm1

“…The result was similar when the prm1D strain was h + and h 2 . The same kind of behavior was found for the dni1D, dni2D, and fus1D mutants (Petersen et al 1995; Clemente-Ramos Figure 7D, and data not shown). The sporulation efficiency of the unilateral WT 3 dni2D prm1D cross was significantly lower than that of the unilateral WT 3 dni2D and WT 3 prm1D crosses ( Figure 7D), suggesting that prm1 + and dni2 + acted in different genetic subpathways for cell fusion.…”

“…The S. cerevisiae FUS1 and the S. pombe fus1 + genes have the same name and both mutants exhibit cell fusion defects during mating, leading to an accumulation of prezygotes (mating intermediates in which the mating partners have established a stable contact but have not yet fused). However, the corresponding proteins are not related; while Saccharomyces cerevisiae Fus1p is a membrane protein (Trueheart et al 1987), S. pombe Fus1p is a formin required for the organization of actin patches at the shmoo tip (Petersen et al 1995(Petersen et al , 1998b. S. pombe diploid nuclei are unstable, such that meiosis occurs immediately after karyogamy, and zygotes give rise to asci containing four ascospores (Nielsen 2004;Yamamoto 2004).…”

“…After agglutination, the cross wall separating both parental cells is degraded, allowing cell fusion. Efficient cell fusion requires the correct organization of the cytoskeleton (Petersen et al 1995(Petersen et al , 1998aKurahashi et al 2002;Doyle et al 2009). The S. cerevisiae FUS1 and the S. pombe fus1 + genes have the same name and both mutants exhibit cell fusion defects during mating, leading to an accumulation of prezygotes (mating intermediates in which the mating partners have established a stable contact but have not yet fused).…”

Cappuccino, a Drosophila maternal effect gene required for polarity of the egg and embryo, is related to the vertebrate limb deformity locus.

“…In addition, there is a 71-amino-acid region within the carboxy-terminal domain that is highly conserved both with the formins and with a number of other proteins (see Fig. 4B (Petersen et al 1995), a genetically undefined S, cerevisiae ORF in GenBank, an Arabidopsis EST, a human EST, and a rice EST. This domain of similarity was independently identified by ourselves and S. Wasserman and has been named the FH2 domain (Castrillon and Wasserman 1994).…”

The Meiotic Bouquet Promotes Homolog Interactions and Restricts Ectopic Recombination inSchizosaccharomyces pombe

“…To identify gene products required for this process, we enriched for mutants of strain GP2640 (h 90 ade6-52 leu1-32 ura4-294 his3-D1 fus1TLEU2 rec12-152TLEU2) as follows. Haploid fus1 mutant cells are unable to mate and therefore cannot sporulate; however, fus1 mutant diploids, if heterozygous at the mating-type locus, are able to properly complete meiosis and sporulation (Petersen et al 1995). By coupling the ability of fus1D to prevent sporulation of haploid cells with the ability to selectively kill nonsporulated cells using glusulase treatment (Ponticelli and Smith 1989), the meiotic progeny of diploid cells can be efficiently selected.…”