Creeping yeast: a simple, cheap and robust protocol for the identification of mating type in Saccharomyces cerevisiae

Abstract

Saccharomyces cerevisiae is an exceptional genetic system, with genetic crosses facilitated by its ability to be maintained in haploid and diploid forms. Such crosses are straightforward if the mating type/ploidy of the strains are known. Several techniques can determine mating type (or ploidy), but all have limitations. Here we validate a simple, cheap and robust method to identify S. cerevisiae mating types. When cells of opposite mating type are mixed in liquid media, they ‘creep’ up the culture vessel sides, a phenotype that can be easily detected visually. In contrast, mixtures of the same mating type or with a diploid simply settle out. The phenotype is observable for several days under a range of routine growth conditions and with different media/strains. Microscopy suggests that cell aggregation during mating is responsible for the phenotype. Yeast knockout collection analysis identified 107 genes required for the creeping phenotype, with these being enriched for mating-specific genes. Surprisingly, the RIM101 signalling pathway was strongly represented. We propose that RIM101 signalling regulates aggregation as part of a wider, previously-unrecognized role in mating. The simplicity and robustness of this method makes it ideal for routine verification of S. cerevisiae mating type, with future studies required to verify its molecular basis.

Cell-surface protein YwfG of Lactococcus lactis binds to α-1,2-linked mannose

“…Non-aggregated yeast cells settle toward the center of round-bottomed vessels and wells, but when aggregated, they assume a uniform distribution across the bottom of the vessel or well. Such a uniform distribution or “creeping” phenotype [ 44 ] is observed in glycosylation-deficient mutants [ 45 ] and in mating yeast cells. Arras et al [ 44 ] have described that yeast cells ‘creep’ up the sides of the culture vessel when cells of opposite mating types are mixed, and they have shown that this ‘creeping’ phenotype is a consequence of cell aggregation.…”

“…Such a uniform distribution or “creeping” phenotype [ 44 ] is observed in glycosylation-deficient mutants [ 45 ] and in mating yeast cells. Arras et al [ 44 ] have described that yeast cells ‘creep’ up the sides of the culture vessel when cells of opposite mating types are mixed, and they have shown that this ‘creeping’ phenotype is a consequence of cell aggregation. The aggregation induced by YwfG 28–336 but not by YwfG 28–270 or YwfG 28–511 was inhibited by the addition of 1 mM d -mannose or α methyl- d -mannoside ( Fig 7B ).…”