Defective Hyphal Development and Avirulence Caused by a Deletion of the SSK1 Response Regulator Gene in Candida albicans

Candida albicans is the most frequently isolated opportunistic fungal pathogen in humans. A number of factors have been associated with the virulence properties of C. albicans, such as adherence to host cells and the ability to undergo the transition from yeast to hyphal growth. This switch is induced in vitro by many environmental conditions, including temperature, pH, and the presence of serum. Diploid strains of Saccharomyces cerevisiae also switch their pattern of growth from unicellular yeast to chains of elongated cells that remain attached to each other (pseudohyphae) under conditions of nitrogen starvation (14). This switch requires STE20, STE11, STE7, KSS1, and STE12, genes of a conserved mitogen-activated protein (MAP) kinase pathway, but also depends on PHD1, a gene which functions in an STE12-independent pathway (3). Based upon functional complementation studies of the STE20, STE7, KSS1, and STE12 genes of S. cerevisiae, the homolog genes CST20 (15, 16), HST7 (15, 16), CEK1 (11), and CPH1 (18) of C. albicans, respectively, have been identified. Mutants of C. albicans containing mutations in each of these genes are unable to undergo the transition from yeast to hyphae on solid media, except when serum is included. In the same way, it was reported that the CPP1 gene, which encodes a phosphatase similar to the MAP kinase phosphatase Msg5p of S. cerevisiae, modulates the activity of the CPH1 pathway, likely by dephosphorylating Cek1p (10). Following a similar experimental approach, the C. albicans genes CaCLA4 (a CLA4 homolog) (17), EFG1 (a PHD1 homolog) (19, 28), CaTUP1 (a TUP1 homolog) (4), and CaRSR1 (an RSR1 homolog) (30) were also studied. The virulence of each of the single mutants described above (except for Δtup1 mutants, with which virulence studies were not performed) was reduced in a murine model of hematogenously disseminated candidiasis (10, 11, 16, 17, 19, 30), but a Δcph1 Δefg1 double mutant was shown to be avirulent (19). However, recent studies indicate that, in fact, the Δcph1 Δefg1 double mutant can still form hyphae when colonizing tissue, indicating that an additional EFG1- and CPH1-independent pathway that regulates morphogenesis in C. albicans may exist (23). More recently, it has been demonstrated that the HOG1 gene of C. albicans, in addition to its role in cytokinesis and response to osmostress in C. albicans (2), also functions in morphogenesis (2). Interestingly, the HOG1 homolog of S. cerevisiae does not function in the filamentation-invasion pathway of S. cerevisiae but in the HOG pathway, which is regulated in part by a two-component cascade whose functional proteins have domains similar to the sensor histidine kinases (Sln1p and Ypd1) and response regulators (Sln1p and Ssk1p) of prokaryotes (3). In this regard, in C. albicans the two-component sensor histidine kinase gene CHK1 (7) seems to regulate the expression of hyphal surface components (5) and perhaps also virulence factors, since a Δchk1 null mutant is avirulent (8), while the NIK1/COS1 gene (21, 27) is required for hyphal development on solid media (1). In previous work, we have identified the putative SSK1 response regulator gene of C. albicans (6). CaSSK1 encodes a protein (CaSsk1p) which is a structural homolog of Ssk1p and Mcs4 from S. cerevisiae and Schizosaccharomyces pombe, respectively. Ssk1p is a response regulator of the two-component cascade that regulates the HOG pathway of S. cerevisiae (20), and Mcs4 is an element of the Sty1 pathway of S. pombe (9, 26). Unlike Ssk1p, which functions only in osmosensing in S. cerevisiae, Mcs4 plays a role in regulating adaptive responses to different stresses, including osmotic and oxidative stresses, and coordinates the responses to these environmental stimuli with the cell cycle (9, 26). Interestingly, we have shown that CaSSK1 fails to complement the lack of SSK1 and mcs4+ in S. cerevisiae and S. pombe, respectively (6). This result indicates that the SSK1 gene of C. albicans may have functions other than modulating the response to osmotic or oxidative stress. In this paper, we present data indicating that SSK1 is involved in the morphogenesis and virulence of C. albicans.