SummaryRho proteins are key regulators of cellular morphogenesis, but their function in filamentous fungi is poorly understood. By generating conditional rho-1 mutants, we dissected the function of the essential GTPase RHO1 in cell polarization and maintenance of cell wall integrity in Neurospora crassa. We identified NCU00668/RGF1 as RHO1-specific exchange factor, which controls actin organization and the cell wall integrity MAK1 MAP kinase pathway through the direct interaction of active RHO1 with the formin BNI1 and PKC1 respectively. The activity of RGF1 is controlled by an intramolecular interaction of its DEP and GEF domains that blocks the activation of the GTPase. Moreover, the N-terminal region including the DEP domain of RGF1 interacts with the plasma membrane sensor NCU06910/WSC1, potentially to activate the cell wall integrity pathway. RHO1 also functions as regulatory subunit of the glucan synthase. N. crassa possesses a second GTPase, RHO2, that is highly homologous to RHO1. RHO2 is of minor importance for growth and does not interact with BNI1. Conditional rho-1;rho-2 double mutants display strong synthetic growth and cell polarity defects. We show that RHO2 does not regulate glucan synthase activity and the actin cytoskeleton, but physically interacts with PKC1 to regulate the cell wall integrity pathway.
In the presented case study, ascomycete fungi and green algae on a marble monument were identified by comparisons of the 18S rRNA gene sequences, which were obtained from DNA either from environmental samples or from enrichment cultures. The organisms were found to be responsible for either black or green surface coverings on different areas of the monument surface. Most fungi were related to plant-inhabiting genera, corresponding to a heavy soiling of the marble surface with honeydew. Whereas green algae of the genera Stichococcus, Chloroidium and Apatococcus were found to be dominant in all samples, isolates of two additional genera were recovered only from enrichment cultures. A reference strain of Apatococcus lobatus and an isolate of Prasiolopsis sp. were investigated with respect to putative surface adhesive structures of the cell envelope. The Prasiolopsis cell walls were covered with a thin adhesive exopolysaccharide layer involved in biofilm formation.
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