An Amyloid Core Sequence in the Major Candida albicans Adhesin Als1p Mediates Cell-Cell Adhesion

Abstract

The human fungal commensal Candida albicans can become a serious opportunistic pathogen in immunocompromised hosts. The C. albicans cell adhesion protein Als1p is a highly expressed member of a large family of paralogous adhesins. Als1p can mediate binding to epithelial and endothelial cells, is upregulated in infections, and is important for biofilm formation. Als1p includes an amyloid-forming sequence at amino acids 325 to 331, identical to the sequence in the paralogs Als5p and Als3p. Therefore, we mutated Val326 to test whether this sequence is important for activity. Wild-type Als1p (Als1pWT) and Als1p with the V326N mutation (Als1pV326N) were expressed at similar levels in a Saccharomyces cerevisiae surface display model. Als1pV326N cells adhered to bovine serum albumin (BSA)-coated beads similarly to Als1pWT cells. However, cells displaying Als1pV326N showed visibly smaller aggregates and did not fluoresce in the presence of the amyloid-binding dye Thioflavin-T. A new analysis tool for single-molecule force spectroscopy-derived surface mapping showed that statistically significant force-dependent Als1p clustering occurred in Als1pWT cells but was absent in Als1pV326N cells. In single-cell force spectroscopy experiments, strong cell-cell adhesion was dependent on an intact amyloid core sequence on both interacting cells. Thus, the major adhesin Als1p interacts through amyloid-like β-aggregation to cluster adhesin molecules in cis on the cell surface as well as in trans to form cell-cell bonds. IMPORTANCE Microbial cell surface adhesins control essential processes such as adhesion, colonization, and biofilm formation. In the opportunistic fungal pathogen Candida albicans, the agglutinin-like sequence (ALS) gene family encodes eight cell surface glycoproteins that mediate adherence to biotic and abiotic surfaces and cell-cell aggregation. Als proteins are critical for commensalism and virulence. Their activities include attachment and invasion of endothelial and epithelial cells, morphogenesis, and formation of biofilms on host tissue and indwelling medical catheters. At the molecular level, Als5p-mediated cell-cell aggregation is dependent on the formation of amyloid-like nanodomains between Als5p-expressing cells. A single-site mutation to valine 326 abolishes cellular aggregation and amyloid formation. Our results show that the binding characteristics of Als1p follow a mechanistic model similar to Als5p, despite its differential expression and biological roles.

A New Function for Amyloid-Like Interactions: Cross-Beta Aggregates of Adhesins form Cell-to-Cell Bonds

“…C. albicans Als1 shares the same sequences of the amyloid core and T domains with Als5, rendering their similarities highly predictable. Experimental studies have indeed confirmed that both adhesins show the same characteristics of cell-to-cell binding initiated by shear-mediated [ 6 , 38 , 43 , 44 , 45 ]. Further, cell-to-cell adhesion occurs with the highest probability and strength when both cells express Als1 with the wild-type cross-β core sequence, while cells that express the non-amyloid substitution V326N have reduced adhesion probability and strength [ 38 ].…”

“…The zipper requires homologous interactions to maintain both the intra- and inter-sheet associations, so sequence identity is key. Thus, these bonds are expected to be strong and highly stable, as observed in assays in vitro [ 6 , 37 , 38 ]. These characteristics were recognized and named ‘SRS adhesions’ for strong reversible specific two decades ago [ 37 ].…”

Adhesion of Staphylococcus aureus to Candida albicans During Co-Infection Promotes Bacterial Dissemination Through the Host Immune Response

“…One of the key players in C. albicans biofilm formation is the Als3 protein ( Zhao et al, 2004 ; Nobile et al, 2006 ), a member of the agglutinin-like sequence ( ALS ) family which is only expressed during the hyphal or pseudo-hyphal phase ( Hoyer et al, 2008 ; Hoyer and Cota, 2016 ). Another ALS family member, ALS1 , shows 81% sequence homology to ALS3 and has overlapping functions but different expression patterns ( Hoyer et al, 1998 ; Liu and Filler, 2011 ; Ho et al, 2019 ). Remarkably, a C. albicans ALS3 deletion strain is only impaired in biofilm formation in vitro but not in vivo , presumably due to the activation of additional adhesin genes, such as ALS1 ( Nobile et al, 2006 ).…”

The dual role of amyloid-β-sheet sequences in the cell surface properties of FLO11-encoded flocculins in Saccharomyces cerevisiae

“…More recently, Lipke, Dufrêne and colleagues developed a new pipette technology called Fluidic Force microscopy enabling to measure forces between single cells [4]. Using this methodology, these authors showed that amyloid-forming sequences have a dual role, namely in the formation of adhesin clusters (nanodomains), which correspond to cis-interactions between molecule of the same cell, and in homophilic adhesion between adhesins on opposing cells (transinteraction) [5,6].…”