Single-cell fluidic force microscopy reveals stress-dependent molecular interactions in yeast mating.

Sexual agglutinins of the budding yeast Saccharomyces cerevisiae are proteins mediating cell aggregation during mating. Complementary agglutinins expressed by cells of opposite mating types "a" and "α" bind together to promote agglutination and facilitate fusion of haploid cells. By means of an innovative single-cell manipulation assay combining fluidic force microscopy with force spectroscopy, we unravel the strength of single specific bonds between a- and α-agglutinins (~100 pN) which require pheromone induction. Prolonged cell-cell contact strongly increases adhesion between mating cells, likely resulting from an increased expression of agglutinins. In addition, we highlight the critical role of disulfide bonds of the a-agglutinin and of histidine residue H273 of α-agglutinin. Most interestingly, we find that mechanical tension enhances the interaction strength, pointing to a model where physical stress induces conformational changes in the agglutinins, from a weak-binding folded state, to a strong-binding extended state. Our single-cell technology shows promises for understanding and controlling the complex mechanism of yeast sexuality.

Fluidic Force Microscopy Captures Amyloid Bonds between Microbial Cells.

Fluidic force microscopy (FluidFM) is a recent force-controlled pipette technology that enables manipulation of single cells. FluidFM can be used for quantification of forces between single cells, and a novel mode of cell-cell adhesion was uncovered: amyloid-like interactions that mediate homophilic adhesion in the fungal pathogen Candida albicans.

Fluidic Force Microscopy Demonstrates That Homophilic Adhesion by Candida albicans Als Proteins Is Mediated by Amyloid Bonds between Cells.

The fungal pathogen Candida albicans frequently forms drug-resistant biofilms in hospital settings and in chronic disease patients. Cell adhesion and biofilm formation involve a family of cell surface Als (agglutinin-like sequence) proteins. It is now well documented that amyloid-like clusters of laterally arranged Als proteins activate cell-cell adhesion under mechanical stress, but whether amyloid-like bonds form between aggregating cells is not known. To address this issue, we measure the forces driving Als5-mediated intercellular adhesion using an innovative fluidic force microscopy platform. Strong cell-cell adhesion is dependent on expression of amyloid-forming Als5 at high cell surface density and is inhibited by a short antiamyloid peptide. Furthermore, there is greatly attenuated binding between cells expressing amyloid-forming Als5 and cells with a nonamyloid form of Als5. Thus, homophilic bonding between Als5 proteins on adhering cells is the major mode of fungal aggregation, rather than protein-ligand interactions. These results point to a model whereby amyloid-like ß-sheet interactions play a dual role in cell-cell adhesion, that is, in formation of adhesin nanoclusters ( cis-interactions) and in homophilic bonding between amyloid sequences on opposing cells ( trans-interactions). Because potential amyloid-forming sequences are found in many microbial adhesins, we speculate that this novel mechanism of amyloid-based homophilic adhesion might be widespread and could represent an interesting target for treating biofilm-associated infections.