Regulation of azole drug susceptibility by Candida albicans protein kinase CK2

Fluconazole resistance of the fungal pathogen Candida albicans can arise through several mechanisms, but the responsible genes and pathways are poorly understood. We report here that mutations in CKA2, identified through an insertional mutagenesis screen, confer fluconazole resistance. CKA2 and its homologue CKA1 specify catalytic subunits of protein kinase CK2. Although cka1 mutations have little effect on fluconazole resistance, CKA1 overexpression suppresses the fluconazole resistance of a cka2 mutant. This observation, along with synthetic cka1-cka2 interactions, argues that Cka1p and Cka2p carry out similar functions. cka2 mutants overexpress CDR1 and CDR2, two fluconazole efflux transporter genes, and a cdr1 mutation decreases resistance of a cka2 mutant, as expected if CDR1 and CDR2 overexpression is responsible for fluconazole resistance of the cka2 mutant. The protein phosphatase calcineurin is required for azole tolerance, and we find that the calcineurin inhibitor cyclosporin reverses fluconazole resistance of cka2 mutants. In addition, a mutation in CRZ1, which specifies a homologue of the Saccharomyces cerevisiae transcription factor that is a major target of calcineurin, suppresses fluconazole resistance of cka2 mutants. Expression analysis of Cka2p-responsive genes argues that Cka2p and Crz1p act through distinct mechanisms. Several clinical fluconazole-resistant isolates overexpress some Cka2p-responsive genes. We suggest that a Cka2p-dependent regulatory pathway is altered by clinically derived azole resistance mutations.