Generation of Viable Candida albicans Mutants Lacking the “Essential” Protein Kinase Snf1 by Inducible Gene Deletion

Abstract

The protein kinase Snf1, a member of the highly conserved AMP-activated protein kinase family, is a central regulator of metabolic adaptation. In the pathogenic yeast <i>Candida albicans</i>, Snf1 is considered to be essential, as previous attempts by different research groups to generate homozygous <i>snf1</i>Δ mutants were unsuccessful. We aimed to elucidate why Snf1 is required for viability in <i>C. albicans</i> by generating <i>snf1</i>Δ null mutants through forced, inducible gene deletion and observing the terminal phenotype before cell death. Unexpectedly, we found that <i>snf1</i>Δ mutants were viable and could grow, albeit very slowly, on rich media containing the preferred carbon source glucose. Growth was improved when the cells were incubated at 37°C instead of 30°C, and this phenotype enabled us to isolate homozygous <i>snf1</i>Δ mutants also by conventional, sequential deletion of both <i>SNF1</i> alleles in a wild-type <i>C. albicans</i> strain. All <i>snf1</i>Δ mutants could grow slowly on glucose but were unable to utilize alternative carbon sources. Our results show that, under optimal conditions, <i>C. albicans</i> can live and grow without Snf1. Furthermore, they demonstrate that inducible gene deletion is a powerful method for assessing gene essentiality in <i>C. albicans</i><b>IMPORTANCE</b> Essential genes are those that are indispensable for the viability and growth of an organism. Previous studies indicated that the protein kinase Snf1, a central regulator of metabolic adaptation, is essential in the pathogenic yeast <i>Candida albicans</i>, because no homozygous <i>snf1</i> deletion mutants of <i>C. albicans</i> wild-type strains could be obtained by standard approaches. In order to investigate the lethal consequences of <i>SNF1</i> deletion, we generated conditional mutants in which <i>SNF1</i> could be deleted by forced, inducible excision from the genome. Unexpectedly, we found that <i>snf1</i> null mutants were viable and could grow slowly under optimal conditions. The growth phenotypes of the <i>snf1</i>Δ mutants explain why such mutants were not recovered in previous attempts. Our study demonstrates that inducible gene deletion is a powerful method for assessing gene essentiality in <i>C. albicans</i>.