title: Electrical phenotypes of calcium transport mutant strains of a filamentous fungus, Neurospora crassa.
abstract: We characterized the electrical phenotypes of mutants with mutations in genes encoding calcium transporters-a mechanosensitive channel homolog (MscS), a Ca(2+)/H(+) exchange protein (cax), and Ca(2+)-ATPases (nca-1, nca-2, nca-3)-as well as those of double mutants (the nca-2 cax, nca-2 nca-3, and nca-3 cax mutants). The electrical characterization used dual impalements to obtain cable-corrected current-voltage measurements. Only two types of mutants (the MscS mutant; the nca-2 mutant and nca-2-containing double mutants) exhibited lower resting potentials. For the nca-2 mutant, on the basis of unchanged conductance and cyanide-induced depolarization of the potential, the cause is attenuated H(+)-ATPase activity. The growth of the nca-2 mutant-containing strains was inhibited by elevated extracellular Ca(2+) levels, indicative of lesions in Ca(2+) homeostasis. However, the net Ca(2+) effluxes of the nca-2 mutant, measured noninvasively with a self-referencing Ca(2+)-selective microelectrode, were similar to those of the wild type. All of the mutants exhibited osmosensitivity similar to that of the wild type (the turgor of the nca-2 mutant was also similar to that of the wild type), suggesting that Ca(2+) signaling does not play a role in osmoregulation. The hyphal tip morphology and tip-localized mitochondria of the nca-2 mutant were similar to those of the wild type, even when the external [Ca(2+)] was elevated. Thus, although Ca(2+) homeostasis is perturbed in the nca-2 mutant (B. J. Bowman et al., Eukaryot. Cell 10:654-661, 2011), the phenotype does not extend to tip growth or to osmoregulation but is revealed by lower H(+)-ATPase activity.
authors.affiliation: Biology Department, York University, Toronto, Ontario, Canada.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Hamam
authors.forename: Roger R
authors.initials: RR
authors.lastname: Lew
mesh_terms: D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002462:Cell Membrane / Q000378:metabolism; D003470:Culture Media / Q000378:metabolism; D046508:Culture Techniques; D003486:Cyanides / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D005800:Genes, Fungal; D006706:Homeostasis; D025301:Hyphae / Q000254:growth & development / Q000502:physiology; D008564:Membrane Potentials; D008839:Microelectrodes; D008928:Mitochondria / Q000502:physiology; D009492:Neurospora crassa / Q000254:growth & development / Q000378:metabolism / Q000502:physiology*; D010641:Phenotype; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism
chemical_list: D003470:Culture Media; D003486:Cyanides; D005656:Fungal Proteins; D025262:Vacuolar Proton-Translocating ATPases; D002118:Calcium