A member of the OSCA/TMEM63 family of mechanosensitive calcium channels participates in cell wall integrity maintenance in Aspergillus nidulans

Abstract

The calF7 mutation in Aspergillus nidulans causes hypersensitivity to the cell wall compromising agents Calcofluor White (CFW) and Congo Red. In this research we demonstrate that the calF7 mutation resides in gene AN2880, encoding a predicted member of the OSCA/TMEM63 family of transmembrane glycoproteins. Those members of the family whose physiological functions have been investigated have been shown to act as mechanosensitive calcium transport channels. Deletion of AN2880 replicates the CFW hypersensitivity phenotype. Separately, we show that CFW hypersensitivity of calF deletion strains can be overcome by inclusion of elevated levels of extracellular calcium ions in the growth medium, and, correspondingly, wild type strains grown in media deficient in calcium ions are no longer resistant to CFW. These observations support a model in which accommodation to at least some forms of cell wall stress is mediated by a calcium ion signaling system in which the AN2880 gene product plays a role. The genetic lesion in calF7 is predicted to result in a glycine-to-arginine substitution at position 638 of the 945-residue CalF protein in a region of the RSN1_7TM domain that is highly conserved amongst filamentous fungi. Homology modeling predicts that the consequence of a G638R substitution is to structurally occlude the principal conductance pore in the protein. GFP-tagged wild type CalF localizes principally to the Spitzenkörper and the plasma membrane at growing tips and forming septa. However, both septation and hyphal morphology appear to be normal in calF7 and AN2880 deletion strains, indicating that any role played by CalF in normal hyphal growth and cytokinesis is dispensable.