Structural basis of flocculin-mediated social behavior in yeast

Abstract

In the budding yeast Saccharomyces cerevisiae, self-recognition and the thereby promoted aggregation of thousands of cells into protective flocs is mediated by a family of cell-surface adhesins, the flocculins (Flo). Based on this social behavior FLO genes fulfill the definition of "greenbeard" genes, which direct cooperation toward other carriers of the same gene. The process of flocculation plays an eminent role in the food industry for the production of beer and wine. However, the precise mode of flocculin-mediated surface recognition and the exact structure of cognate ligands have remained elusive. Here, we present structures of the adhesion domain of a flocculin complexed to its cognate ligands derived from yeast high-mannose oligosaccharides at resolutions up to 0.95 Å. Besides a PA14-like architecture, the Flo5A domain reveals a previously undescribed lectin fold that utilizes a unique DcisD calcium-binding motif for carbohydrate binding and that is widely spread among pro-and eukaryotes. Given the high abundance of high-mannose oligosaccharides in yeast cell walls, the Flo5A structure suggests a model for recognition, where social non-selfinstead of unsocial self-interactions are favored.altruism | molecular recognition | fungal development | atomic resolution S elf-recognition is key to both microbial growth and tissue formation. The molecular basis of this phenomenon is well understood only for a few cases-e.g., during neural differentiation and immune recognition. In the microbial world, the formation of multicellular structures by self-recognition is mediated by specific, surface-exposed adhesins and provides protection and promotes long-term survival, substrate exploration, or host invasion (1, 2). In the budding yeast Saccharomyces cerevisiae, aggregation of vegetative cells is a paradigm for biofilm formation (3, 4) and the evolution of social behavior (5, 6). Self-recognition in this organism is mediated by flocculin proteins, which belong to a family of fungal adhesins that are present in pathogenic yeasts as well (7). The general architecture of flocculins includes an N-terminal A domain corresponding to a lectin-like adhesin domain, a stalk-like, repetitive and highly glycosylated B domain, and a C domain that carries a GPI anchoring site (8). The genome of S. cerevisiae contains at least four functional FLO genes (FLO1, FLO5, FLO9, and FLO10) whose N-terminal A domains are predicted to belong to the PA14-like protein family (9) first described in the anthrax-protective antigen (10). This protein family has been found to be widely distributed among all domains of life including human galactosyltransferases and fibrocystin (9, 11).Flocculins confer dominant, calcium-dependent cell-cell adhesion that can be inhibited by hexoses like mannose (12, 13). This process is known as flocculation and has been exploited for centuries by the brewing industry (14). Flocculin genes also mediate social behavior of S. cerevisiae and have here been found to fulfill the definition of "greenbeard" genes (5, 6). E...

Flocculation in ale brewing strains of Saccharomyces cerevisiae: re-evaluation of the role of cell surface charge and hydrophobicity

“…These data strongly suggest that the concomitant increase of CSH and the onset of flocculation, in ale brewing yeast cells, can now be explained by the expression of flocculins on yeast cells surface. This possibility is in agreement with the fact that N-and C-terminal regions of Flo1 protein are more hydrophobic than the rest of the protein (Watari et al 1994); N-terminal region corresponds to the domain A of Flo proteins, which is responsible by the recognition and adhesion to receptors (Kobayashi et al 1998;Veelders et al 2010). …”

“…The N-terminal region of Flo1 protein (domain A) contains the sugar recognition domain, which is important for flocculation definition of Flo1 and NewFlo phenotypes (Kobayashi et al 1998). A detailed analysis of the adhesion domain (domain A) of the flocculin Flo5 revealed a bipartite organization constituted by a large β-sandwich domain (which is related with the PA14 domain of the anthrax-protective antigen); this domain takes an insertion, the Flo5 subdomain, which is composed by five short β-strands, stabilized by two disulphide bridges (Rigden et al 2004;Veelders et al 2010). It was also shown that calcium is directly involved in carbohydrate binding and not indirectly in the stabilization of the rodlike B domain (Veelders et al 2010).…”

125th Anniversary Review: Yeast Flocculation and Sedimentation in Brewing

“…It has been suggested that Ca 2+ binds to flocculin proteins and is necessary for flocculin to obtain the correct structural conformation 51 . A recent crystallization and structural study of flocculins showed that Ca 2+ is directly involved in carbohydrate binding 101 . Flocculation is a reversible phenomenon, so that cell flocs can be dissociated, e.g., by adding a chelating agent that removes calcium ions or by adding mannose, which competitively displaces cell wall mannose residues from flocculin binding sites.…”

Cleanup of industrial effluents containing heavy metals: a new opportunity of valorising the biomass produced by brewing industry

“…According to current knowledge, the flocculation of brewing yeasts occurs due to the interaction of a specific lectin-like protein (only present on flocculent cells) with the surface carbohydrate residues of the α-mannan (receptors) of neighbouring cells (Soares 2011). It was described that Ca ions are directly involved in the binding of the flocculation lectin to the α-mannan receptors (Veelders et al 2010). Yeast flocculation is influenced by characteristics of the medium and the presence of specific FLO genes (such as FLO1, FLO5,FLO8,FLO9,and FLO10) (Soares 2011).…”