Molecular Mechanism of Flocculation Self-Recognition in Yeast and Its Role in Mating and Survival

Abstract

We studied the flocculation mechanism at the molecular level by determining the atomic structures of N-Flo1p and N-Lg-Flo1p in complex with their ligands. We show that they have similar ligand binding mechanisms but distinct carbohydrate specificities and affinities, which are determined by the compactness of the binding site. We characterized the glycans of Flo1p and their role in this binding process and demonstrate that glycan-glycan interactions significantly contribute to the cell-cell adhesion mechanism. Therefore, the extended flocculation mechanism is based on the self-interaction of Flo proteins and this interaction is established in two stages, involving both glycan-glycan and protein-glycan interactions. The crucial role of calcium in both types of interaction was demonstrated: Ca2+ takes part in the binding of the carbohydrate to the protein, and the glycans aggregate only in the presence of Ca2+. These results unify the generally accepted lectin hypothesis with the historically first-proposed “Ca2+-bridge” hypothesis. Additionally, a new role of cell flocculation is demonstrated; i.e., flocculation is linked to cell conjugation and mating, and survival chances consequently increase significantly by spore formation and by introduction of genetic variability. The role of Flo1p in mating was demonstrated by showing that mating efficiency is increased when cells flocculate and by differential transcriptome analysis of flocculating versus nonflocculating cells in a low-shear environment (microgravity). The results show that a multicellular clump (floc) provides a uniquely organized multicellular ultrastructure that provides a suitable microenvironment to induce and perform cell conjugation and mating.

Flocculation of Saccharomyces cerevisiae is dependent on activation of Slt2 and Rlm1 regulated by the cell wall integrity pathway

“…Thus, these results suggest that the activation of the CWI pathway ( MSG5 deletion, CFW or heat stress) increases the basal levels of Slt2 phosphorylation resulting in FLO gene expression. Therefore, we conclude that the activation of the CWI pathway is essential for eliciting the expression of FLO genes and flocculation phenotype which provides a suitable microenvironment required for cell survival under stress conditions (Goossens et al ., ).…”

“…The genetic, epigenetic and environmental factors; mutations in certain genes, nutritional stress, temperature, pH, oxygen supply and alteration in cell wall composition have been shown to induce flocculation phenotypes (Soares, 2011;Kim and Rose, 2015;Liu et al, 2015). Flocculins are lectin-like cell wall proteins that bind to mannose residues on the cell wall of neighboring cells leading to flocculation (Van Mulders et al, 2009, Goossens et al, 2015. Yeast subtelomeric loci carry five dominant FLO genes; FLO1, FLO5, FLO9, FLO10 and FLO11, all of which are epigenetically regulated (Halme et al, 2004) and responsible for flocculation and biofilm formation (Smukalla et al, 2008;Goossens et al, 2011;Sim et al, 2013).…”

Expression of AtMed15 of Arabidopsis in yeast causes flocculation and increases ethanol production in yeast culture

“…Cations play very important role in yeast flocculation; mostly they promote flocculation. The free and labile Ca 2+ ions in the media enable cell wall bound lectins/mannoproteins adopt their active conformation, which in turn interacts with carbohydrate residues of neighbouring cells to cause flocculation3536. In order to see effect of cations, EDTA was added in the culture media.…”

“…Expression of Aga1 and Flo11/Muc1 might also contribute to the robustness of AtMed15 conferred flocculation in yeast culture. The Lectin theory of flocculation explains that in the presence of calcium, cell wall bound lectins/flocculins bind to mannose moieties of cell wall of neighbouring cell to form flocs353638. The AtMed15-induced flocculation showed maximum sensitivity to the mannose as compared to glucose, sucrose, maltose and fructose (Fig.…”

Dominance of wine Saccharomyces cerevisiae strains over S. kudriavzevii in industrial fermentation competitions is related to an acceleration of nutrient uptake and utilization

“…Indeed, some of the genes we found as differentially expressed in competition, which functions remain unknown, could be specifically involved in microbial interactions. Among them, the FLO gene family could be a clear candidate as the main function of these genes is self-recognition and flocculation interaction with other cells (Goossens et al, 2015). In fact, a study in which FLO1, FLO5, FLO9 and FLO10 expression was controlled in co-cultures of S. cerevisiae and several non-Saccharomyces yeast, aggregation-flocculation and yeast competitive fitness varied depending on the competitor species and the overexpressed flocculin, which implies a species-or strainspecific mechanism of cell-to-cell interaction (Rossouw et al, 2015).…”