Single allele knock-out of Candida albicans CGT1 leads to unexpected resistance to hygromycin B and elevated temperature

Almost all eukaryotic mRNAs are capped at their 5'-terminus. Capping is cru cial for stability, processing, nuclear export and efficient translation of mRNA, We studied the phenotypic effects elicited by depleting a Candida al bicans strain of mRNA 5'-guanylyltransferase (mRNA capping enzyme; CGT1). C onstruction of a Cgt1-deficient mutant was achieved by URA-blaster-mediated genetic disruption of one allele of the CGT1 gene, which was localized on chromosome III. The resulting heterozygous mutant exhibited an aberrant col ony morphology resembling the 'irregular wrinkle' phenotype typically obtai ned from a normal C. albicans strain upon mild UV treatment, Its level of C GT1 mRNA was reduced two- to fivefold compared to the parental strain. Prot eome analysis revealed a large number of differentially expressed proteins confirming the expected pleiotropic effect of CGT1 disruption. The disrupte d strain was significantly more resistant to hygromycin B, an antibiotic wh ich decreases translational fidelity, and showed increased resistance to he at stress. Proteome analysis revealed a 50-fold overexpression of Ef-1 alph a p and a more than sevenfold overexpression of the cell-wall heat-shock pr otein Ssa2p. Compared to a reference strain, the cgt1/CGT1 heterozygote was equally virulent for mice and guinea pigs when tested in an intravenous in fection model of disseminated candidiasis.