CRR1, a gene encoding a putative transglycosidase, is required for proper spore wall assembly in Saccharomyces cerevisiae

Abstract

In Saccharomyces cerevisiae, sporulation is a developmental process that converts a single cell into four haploid spores. The four haploid nuclei are encapsulated within multilayered spore walls that protect them against stressful conditions. The formation of the spore-specific cell wall is a highly coordinated process that requires the participation of enzymic activities for biosynthesis, degradation, and cross-linking between components. Here the sporulation-specific gene CRR1, encoding a putative transglycosidase that is required for proper spore wall assembly, is described. Both the transcription of CRR1 and the synthesis of Crr1p were induced biphasically under sporulating conditions, with a first expression peak displaying kinetics similar to those of the middle to middle-late sporulation-specific genes, and a second late peak after 24 h under these conditions. Localization studies revealed that Crr1p localized to the spore wall that surrounds each of the four ascospores within the mature asci. Mutation of this gene had no effect on the efficiency of spore formation. However, crr1 mutant spores were sensitive to hydrolytic enzymes such as glusulase and to heat-shock treatments, underscoring the importance of this gene in the proper formation and assembly of the ascospore wall. Moreover, the deletion of CRR1 had additive effects with respect to the sensitivity of cda1 cda2 mutants to these treatments. Interestingly, overexpression of CRR1 not only complemented the phenotype of the crr1 strain but also rendered spores more resistant to the stress conditions than the wild-type. Like other mutants impaired in the formation of the spore outer layer, crr1 mutants were permeable to Calcofluor White. Finally, detailed analysis by electron microscopy of the spore walls in the crr1 mutants revealed a defect in the assembly of the spore wall components, suggesting a role for Crr1p in the cross-linking between the inner (glucan/mannoprotein) and the outer (chitosan/dityrosine) spore layers.

A Barley Xyloglucan Xyloglucosyl Transferase Covalently Links Xyloglucan, Cellulosic Substrates, and (1,3;1,4)-β-D-Glucans

“…Emerging information on the remodeling of fungal cell walls during spore formation and under stress indicates that glycosylphosphatidylinositol-anchored transferase enzymes, some of which are members of family GH16, might also be involved in linking different polysaccharides such as ␤-D-glucans and chitin in the wall (45). There are also indications that pectic polysaccharides might be covalently linked with xyloglucans in plant cell walls (10 -13).…”

The Smk1p MAP kinase negatively regulates Gsc2p, a 1,3-β-glucan synthase, during spore wall morphogenesis in Saccharomyces cerevisiae

“…Several genes required for the proper production of the outer spore wall layers have been identified, and include SSP2, CRR1, SPO77, OSW1, and MUM3 (2,26,27). These genes may be important in working with or in parallel to SMK1 to regulate the transition between inner and outer spore wall deposition.…”

Architecture and Biosynthesis of the Saccharomyces cerevisiae Cell Wall

“…Ascopores from Crr1-deficient diploids show increased sensitivity to heat shock and lytic enzymes, and these defects are exacerbated when the chitin deacetylases Cda1 and Cda2 are also absent. These findings suggest a role for Crr1 in generating cross-links between the b-glucan and chitosan or chitin during ascospore wall maturation (Gómez-Esquer et al 2004). …”

“…Crr1-GFP fusions are localized on the surface of ascospores, and homozygous crr1D diploids have ascospore wall abnormalities, with irregular deposition of the outer dityrosine and chitosan layers over the inner b-glucan layer (Gómez-Esquer et al 2004). Ascopores from Crr1-deficient diploids show increased sensitivity to heat shock and lytic enzymes, and these defects are exacerbated when the chitin deacetylases Cda1 and Cda2 are also absent.…”