Possible additional roles in mating for Ustilago maydis Rho1 and 14-3-3 homologues

Abstract

Both the Rho GTPases and 14-3-3 proteins each belong to ubiquitous families of proteins involved in a variety of cellular processes, including cytokinesis, cell polarity, cellular differentiation and apoptosis. In fungi, these components of signaling pathways are involved in cell cycle regulation, cytokinesis and virulence. We study cellular differentiation and pathogenesis for Ustilago maydis, the dimorphic fungal pathogen of maize. We have reported on the interactions of Pdc1, a U. maydis homologue of human 14-3-3varepsilon, with Rho1, a small GTP binding protein; these proteins participate in cell polarity and filamentation pathways that include another small G protein, Rac1, and its effector PAK kinase, Cla4. Here we describe additional experiments that explore possible relationships of Pdc1 and Rho1 with another PAK-like kinase pathway and with the a matingtype locus.

Keywords: MAPK pathway; cell cycle; cell polarity; cytokinesis; difopein; filamentation; mating and pheromone response.

Figures 1

Inhibition of Pdc1 activity by the synthetic peptide inhibitor, difopein, leads to elongated cells. The Pcrg1-controlled difopein expression-construct is integrated into the genome at the ip locus. Arabinose was used for the induction of difopein production and this leads to cell elongation as Pdc1 is inhibited. In contrast, in dextrose, when difopein is not expressed, cells appear like wild type cells growing in arabinose-containing media. Scale bar = 10 µm.

Figures 2

Overexpression of rho1 or pdc1 in the absence of Smu1 leads to multiple-bud morphology. Deletion of smu1 does not cause any discernable phenotype (Δsmu1). Interestingly, when we overexpressed either pdc1 or rho1 in the smu1 background, the cells appeared as aggregates. The promoters of pdc1 and rho1 were replaced with the Pcrg1, so that they would be overexpressed in the presence of arabinose.

Figures 3

Overexpression of pdc1 in SG200 cells leads to cell separation defect. SG200 cells are typically cigar-shaped. However, when we overexpressed pdc1 in this cell line, the cells appear highly branched. The promoter of pdc1 was replaced with the Pcrg1. In arabinose-containing media the gene would be overexpressed.