A conserved fungal hub protein involved in adhesion and drug resistance in the human pathogen Candida albicans

Abstract

Drug resistance and cellular adhesion are two key elements of both dissemination and prevalence of the human fungal pathogen Candida albicans . Smi1 belongs to a family of hub proteins conserved among the fungal kingdom whose functions in cellular signaling affect morphogenesis, cell wall synthesis and stress resistance. The data presented here indicate that C. albicans SMI1 is a functional homolog of Saccharomyces cerevisiae KNR4 and is involved in the regulation of cell wall synthesis. Expression of SMI1 in S. cerevisiae knr4 Δ null mutants rescued their sensitivity to caspofungin and to heat stress. Deletion of SMI1 in C. albicans resulted in sensitivity to the cell-wall-perturbing compounds Calcofluor White and Caspofungin. Analysis of wild-type and mutant cells by Atomic Force Microscopy showed that the Young’s Modulus (stiffness) of the cell wall was reduced by 85% upon deletion of SMI1 , while cell surface adhesion measured by Force Spectroscopy showed that the surface expression of adhesive molecules was also reduced in the mutant. Over-expression of SMI1 , on the contrary, increased cell surface adhesion by 6-fold vs the control strain. Finally, Smi1-GFP localized as cytoplasmic patches and concentrated spots at the sites of new cell wall synthesis including the tips of growing hyphae, consistent with a role in cell wall regulation. Thus, Smi1 function appears to be conserved across fungi, including the yeast S. cerevisiae, the yeast and hyphal forms of C. albicans and the filamentous fungus Neurospora crassa .

Saccharomyces cerevisiae CellWall Remodeling in the Absence of Knr4 and Kre6 Revealed by Nano-FourierTransform Infrared Spectroscopy

“…[13][14][15][16] In the human fungal pathogen C. albicans, Knr4 has two homologs, Smi1 and Smi1B, that are involved in adhesion and antibiotic resistance. 17,18 These observations show that Kre6, Knr4, and their homologs have conserved roles in cell wall biogenesis among these fungi.…”

“…In the studies of Kre6 8 – 12 and Knr4 13 – 18 above, yeasts and wall composition were investigated by conventional fluorescence staining of cells or by extraction and isolation of components for chemical analysis. Infrared (IR) spectroscopy has been used to gain insight into cellular composition based on known IR absorption bands, many of which are distinct for various biocomponents: proteins, fats, and sugars, including chitin and glucans.…”

Adhesins in the virulence of opportunistic fungal pathogens of human

“…Strain that overexpress this protein has been found to have greater cell wall adhesion by many folds than the wild type strain. It is also involved in hyphal morphogenesis (Martin-Yken et al 2018 ). In C. albicans , the EAP1 gene (a putative cell-wall adhesin) has been identified, and in silico analysis indicated that it consists a signal peptide, a GPI-anchor site and has homology with various genes of many yeast encoding CWPs.…”

The Conserved Yeast Protein Knr4 Involved in Cell Wall Integrity is a Multi-Domain Intrinsically Disordered Protein

“…20,26,[28][29][30] Interestingly, KNR4 orthologs in C. albicans, SMI1 and SMI1B, are involved in pathogenesis and in biofilm resistance to antifungal agents. [31][32] Knr4 has two experimentally-confirmed properties out of the three required for a hub protein: it is at the crossroad of essential cellular pathways and it has a plethora of partners in the yeast PPI network. The third property required for Knr4 to be a hub in PPI networks is intrinsic structural flexibility.…”

“…In eukaryotes, Knr4/Smi1 proteins are specific to the fungal kingdom and their deletion in the yeasts S. cerevisiae and C. albicans leads to hypersensitivity to certain antifungal agents. 11,[31][32] Targeting Knr4 interaction regions could lead to promising compounds that could be used in combinatorial therapies to reduce the high doses of antifungal agents currently required to control human pathogenic fungi. 37…”