The crk1 gene encodes an Ime2‐related protein that is required for morphogenesis in the plant pathogen Ustilago maydis

Abstract

SummaryThe fungal pathogen Ustilago maydis alternates between budding and filamentous growth during its life cycle. This dimorphic transition is regulated by environmental factors and mating. We cloned a new gene, crk1 , which encodes a protein with sequence similarity to Ime2, a kinase involved in developmental choices in S. cerevisiae . Disruption of the crk1 gene in U. maydis resulted in cells that are unable to respond in an appropriate manner to environmental stimuli and show defects in morphogenesis and cell cycle adjustment to changing conditions. We have analysed the regulation of the crk1 gene and demonstrated that cAMP and MAPK pathways have opposite influences on the transcript levels for crk1 . Furthermore, we have shown that alterations in the components of these pathways impair the ability of the cellular machinery to adapt to changing conditions. These results demonstrate an important role for the crk1-encoded protein in the morphogenesis and environmental adaptation in Ustilago maydis .

The distinct wiring between cell cycle regulation and the widely conserved Morphogenesis-Related (MOR) pathway in the fungus Ustilago maydis determines the morphological outcome

“…One of these elements appears to be the kinase Crk1. This MAPK-like kinase, which has roles during the mating process in U. maydis (Garrido and Pérez-Martín, 2003;Garrido et al, 2004), had strong effects on the ability of cells to sustain polar growth: cells defective in this kinase had a rounder morphology that contrasted with the elongated bud morphology of wild-type U. maydis cells, while overexpression of crk1 resulted in a dramatic elongated growth (supplementary material S6). In cells arrested in G2 phase after impairment of mitotic CDK activity, crk1 expression level dramatically raised (supplementary material Fig.…”

Inhibitory phosphorylation of a mitotic cyclin-dependent kinase regulates the morphogenesis, cell size and virulence of the smut fungusUstilago maydis

“…DNA, RNA and protein analysis U. maydis DNA and RNA isolation, the preparation of protein extracts, northern and western blotting, and immunoprecipitations were all performed as described previously (Tsukuda et al, 1988;Garrido and Pérez-Martín, 2003;García-Muse et al, 2004;Garrido et al, 2004). The anti-PSTAIRE (Santa Cruz Biotechnology), anti-myc 9E10, and anti-VSV (Roche Diagnostics Gmb) antibodies were all used at a dilution of 1:10,000 in phosphate-buffered saline + 0.1% Tween + 10% dry milk.…”

A member of the Fizzy-related family of APC activators is regulated by cAMP and is required at different stages of plant infection byUstilago maydis

“…3C). Enlargement of the G1 phase is part of the adaptive response to decreasing nutritional condition in U. maydis wild-type cells (Garrido and Pérez-Martín, 2003 G1, thus ensuring a proper G1 length (Blanco et al, 2000;Yamaguchi et al, 2000). In U. maydis, Clb1, which is a target of Cru1, is involved in S-phase promotion (Garcia-Muse et al, 2004).…”

“…U. maydis DNA isolation and transformation was performed as previously described (Tsukuda et al, 1988). RNA isolation and northern analysis were performed as described previously (Garrido and Pérez-Martín, 2003). For mfa1 probe a 0.67-kb EcoRV fragment was used as described previously .…”

“…Image processing was performed with Photoshop (Adobe). Nuclear staining was done using DAPI staining as described previously (Garrido and Pérez-Martín, 2003). WGA staining was performed as described .…”