Saccharomyces cerevisiae YCRO17c/CWH43 encodes a putative sensor/transporter protein upstream of the BCK2 branch of the PKC1‐dependent cell wall integrity pathway

Abstract

The Saccharomyces cerevisiae cwh43-2 mutant, originally isolated for its Calcofluor white hypersensitivity, displays several cell wall defects similar to mutants in the PKC1-MPK1 pathway, including a growth defect and increased release of b-1,6-glucan and bglucosylated proteins into the growth medium at increased temperatures. The cloning of CWH43 showed that it corresponds to YCR017c and encodes a protein with 14-16 transmembrane segments containing several putative phosphorylation and glycosylation sites. The N-terminal part of the amino acid sequence of Cwh43p shows 40% similarity with the mammalian FRAG1, a membrane protein that activates the fibroblast growth factor receptor of rat osteosarcoma (FGFR2-ROS) and with protein sequences of four uncharacterized ORFs from Caenorhabditis elegans and one from Drosophila melanogaster. The C-terminus of Cwh43p shows low similarities with a xylose permease of Bacillus megaterium and with putative sugar transporter from D. melanogaster, and has 52% similarity with a protein sequence from a Schizosaccharomyces pombe cDNA. A Cwh43-GFP fusion protein suggested a plasma membrane localization, although localization to the internal structure of the cells could not be excluded, and it concentrates to the bud tip of small budded cells and to the neck of dividing cells. Deletion of CWH43 resulted in cell wall defects less pronounced than those of the cwh43-2 mutant. This allelespecific phenotype appears to be due to a G-R substitution at position 57 in a highly conserved region of the protein. Genetic analysis places CWH43 upstream of the BCK2 branch of the PKC1 signalling pathway, since cwh43 mutations were synthetic lethal with pkc1 deletion, whereas the cwh43 defects could be rescued by overexpression of BCK2 and not by high-copy-number expression of genes encoding downstream proteins of the PKC1 pathway. However, unlike BCK2, whose disruption in a cln3 mutant resulted in growth arrest in G 1 , no growth defect was observed in a double cwh43 cln3 mutants. Taken together, it is proposed that CWH43 encodes a protein with putative sensor and transporter domains acting in parallel to the main PKC1-dependent cell wall integrity pathway, and that this gene has evolved into two distinct genes in higher eukaryotes.

Putative Stress Sensors WscA and WscB Are Involved in Hypo-Osmotic and Acidic pH Stress Tolerance in Aspergillus nidulans

“…Cwh43 is a putative sensor/ transporter protein that acts upstream of the BCK2 branch of the PKC1-dependent cell wall integrity pathway and is involved in cell wall biogenesis (29). The principal structural feature of Cwh43 is the presence of 14 to 16 transmembrane segments and several putative glycosylation and phosphorylation sites.…”

Structure–function analysis of Knr4/Smi1, a newly member of intrinsically disordered proteins family, indispensable in the absence of a functional PKC1–SLT2 pathway in Saccharomyces cerevisiae

“…Through a multicopy gene expression strategy, we have not been able to find any multicopy suppressor that could overcome the synthetic growth defect of a knr4∆bck1∆ or knr4∆slt2∆. Also overexpression of BCK2, which encodes a protein known to interact and to share partially redundant functions with Knr4 (Martin-Yken et al, 2001;Basmaji et al, 2006) did not rescue growth in these double mutants (data not shown). Altogether, and assuming that all yeast genes were covered in the suppression experiments, these genetic data indicate that there is no other protein that can perform a redundant function with Knr4 in this context.…”

Plant-derived antifungal agent poacic acid targets β-1,3-glucan

“…Among the top 10 deletion mutants sensitive to poacic acid, we detected enrichment for genes involved in the gene ontology (GO) category fungal-type cell wall organization (P < 0.01). These mutants included deletion alleles of BCK1 (bypass of C kinase), which encodes an MAPKKK in the Pkc1p (protein kinase C) cell wall integrity signaling pathway (PKC pathway); CWH43, which encodes a membrane protein involved in cell wall biogenesis and its null mutation is synthetically lethal with PKC1 mutants (29); ROM2 (Rho1 multicopy suppressor), a GDP/GTP exchange factor for Rho1p and another component of the PKC pathway; and ACK1, which seems to encode an upstream activator of Pkc1p and has a physical interaction with Rom2p. Overall, the PKC pathway was the most sensitive pathway to poacic acid (Pathway z score = −7.85) (Fig.…”