Using Genomics to Shape the Definition of the Agglutinin-Like Sequence (ALS) Family in the Saccharomycetales

Abstract

The Candida albicans agglutinin-like sequence (ALS) family is studied because of its contribution to cell adhesion, fungal colonization, and polymicrobial biofilm formation. The goal of this work was to derive an accurate census and sequence for ALS genes in pathogenic yeasts and other closely related species, while probing the boundaries of the ALS family within the Order Saccharomycetales. Bioinformatic methods were combined with laboratory experimentation to characterize 47 novel ALS loci from 8 fungal species. AlphaFold predictions suggested the presence of a conserved N-terminal adhesive domain (NT-Als) structure in all Als proteins reported to date, as well as in S. cerevisiae alpha-agglutinin (Sag1). Lodderomyces elongisporus, Meyerozyma guilliermondii, and Scheffersomyces stipitis were notable because each species had genes with C. albicans ALS features, as well as at least one that encoded a Sag1-like protein. Detection of recombination events between the ALS family and gene families encoding other cell-surface proteins such as Iff/Hyr and Flo suggest widespread domain swapping with the potential to create cell-surface diversity among yeast species. Results from the analysis also revealed subtelomeric ALS genes, ALS pseudogenes, and the potential for yeast species to secrete their own soluble adhesion inhibitors. Information presented here supports the inclusion of SAG1 in the ALS family and yields many experimental hypotheses to pursue to further reveal the nature of the ALS family.

Development and Use of a Monoclonal Antibody Specific for the Candida albicans Cell-Surface Protein Hwp1

“…The Hwp1 Gln-Pro-rich sequences are present in multiple repeated copies ( Staab et al., 1999 ), presumably each with potential to participate in adhesive interactions with BECs. Other C. albicans repeat-containing adhesins can have often-extreme allelic variation ( Zhang et al., 2003 ; Hoyer et al., 2008 ; Oh et al., 2021 ), leading to questions about diversity of HWP1 alleles across C. albicans isolates. Analysis of HWP1 restriction fragment length polymorphisms was proposed as a diagnostic assay to distinguish C. albicans from C. dubliniensis , suggesting stability of fragment size, and therefore repeat copy number across isolates in the same species.…”

“…Aga2 is disulfide bonded to the Aga1 anchor protein that displays Aga2 on the cell surface. The idea that Als proteins are orthologs of S. cerevisiae alpha-agglutinin has gained support as the number of genomic sequences continues to grow ( Oh et al., 2021 ). For example, CGOB predicts that S. cerevisiae SAG1 is syntenic with a C. dubliniensis ALS locus.…”

“…For example, CGOB predicts that S. cerevisiae SAG1 is syntenic with a C. dubliniensis ALS locus. Also, Sag1 and Als proteins appear to share a similar structure of the N-terminal domain that is responsible for adhesive (ligand-binding) function ( Oh et al., 2021 ). In contrast, the idea that Hwp1 is an ortholog of S. cerevisiae a -agglutinin (Aga1) seems prompted by the observation that Hwp1 production and localization during C. albicans mating is specific to the a/a portion of the conjugation bridge ( Daniels et al., 2003 ).…”

Development and validation of monoclonal antibodies specific for Candida albicans Als2, Als9-1, and Als9-2

“…Agglutinin-like sequence (Als) proteins are large, cell-surface glycoproteins that function in adhesive and aggregative interactions [reviewed in 1]. Candida albicans has eight distinct ALS loci (ALS1 to ALS7, ALS9); many species in the Saccharomycetales have at least one ALS gene [2]. Als proteins have a secretory signal peptide at the N-terminal end and a GPI anchor addition site at the C-terminal end that direct their processing and modification via the secretory pathway to a final localization linked to ß-1,6-glucan in the fungal cell wall [3].…”

“…Als aggregative interactions are mediated by an amyloid-forming region that is part of the N-terminal domain [6]. Although the general NT-domain structure appears to be conserved among Als proteins across a wide variety of fungal species [2], sufficient sequence variation exists to use the NT domain to raise monoclonal antibodies (MAbs) that specifically recognize individual Als proteins.…”