Candida albicans INT1-induced filamentation in Saccharomyces cerevisiae depends on Sla2p

Abstract

The Candida albicans INT1 gene is important for hyphal morphogenesis, adherence, and virulence (C. Gale, C. Bendel, M. McClellan, M. Hauser, J. M. Becker, J. Berman, and M. Hostetter, Science 279:1355-1358, 1998). The ability to switch between yeast and hyphal morphologies is an important virulence factor in this fungal pathogen. When INT1 is expressed in Saccharomyces cerevisiae, cells grow with a filamentous morphology that we exploited to gain insights into how C. albicans regulates hyphal growth. In S. cerevisiae, INT1-induced filamentous growth was affected by a small subset of actin mutations and a limited set of actin-interacting proteins including Sla2p, an S. cerevisiae protein with similarity in its C terminus to mouse talin. Interestingly, while SLA2 was required for INT1-induced filamentous growth, it was not required for polarized growth in response to several other conditions, suggesting that Sla2p is not required for polarized growth per se. The morphogenesis checkpoint, mediated by Swe1p, contributes to INT1-induced filamentous growth; however, epistasis analysis suggests that Sla2p and Swe1p contribute to INT1-induced filamentous growth through independent pathways. The C. albicans SLA2 homolog (CaSLA2) complements S. cerevisiae sla2Delta mutants for growth at 37 degrees C and INT1-induced filamentous growth. Furthermore, in a C. albicans Casla2/Casla2 strain, hyphal growth did not occur in response to either nutrient deprivation or to potent stimuli, such as mammalian serum. Thus, through analysis of INT1-induced filamentous growth in S. cerevisiae, we have identified a C. albicans gene, SLA2, that is required for hyphal growth in C. albicans.

FIG. 1

Actin is highly polarized in S. cerevisiae cells expressing INT1. DIC images (A) and fluorescence images (B and C) of rhodamine-phalloidin-stained strain YJB2603 cells expressing INT1 (A and B) after 12 h of growth on galactose. (C) Rhodamine-phalloidin-stained YJB2603 cells were grown on glucose (to repress INT1 expression).

FIG. 2

INT1-induced filamentous growth is reduced by specific actin mutations. Shown are DIC micrographs of strains carrying the indicated ACT1 alleles (ACT1, YJB2603; act1-101, YJB2604; act1-104, YJB2614; act1-124, YJB2610; act1-129, YJB2611; act1-120, YJB2608) and expressing INT1 after growth on galactose for 16 h. Strains are listed in Tables 1 and 2.

FIG. 3

INT1-induced filamentous growth requires SLA2, but not ABP1 or SAC6. Shown are DIC micrographs of isogenic wild-type and mutant strains with the relevant genotype indicated. Strains are listed in Tables 1 and 3.

FIG. 4

SLA2 is not required for all types of polarized growth. (A) DIC micrographs of strains carrying the cdc3-6 allele, which results in polarized growth in the presence (YJB5565; left) or absence (YJB5566; right) of SLA2. (B) DIC micrographs of MATa wild-type (YJB2489) and sla2Δ (YJB4786) strains exposed to 300 μg of α-factor/ml.

FIG. 5

SLA2 and SWE1 contribute independently to INT1-induced filamentous growth. The percentage of INT1-induced filaments was determined by spreading cells onto plates containing 2% galactose and counting the cells producing filaments and the total number of cells on the plate 18 h after plating. Two isolates transformed with pGAL-INT1 were used for each experiment, and a minimum of 200 cells was counted for each strain. The wild-type strain produced 95% filamentous cells under these conditions.

FIG. 6

S. cerevisiae SLA2 and C. albicans SLA2 have a talin-like domain which is not required for Sla2p function in S. cerevisiae. (A) Comparison of ScSla2p, CaSla2p, and mouse talin domains. Amino acid positions of the domains illustrated are noted below the genes. Percent identity between domains of the proteins is indicated. Pro, proline-rich region; Q and L, glutamine- and leucine-rich regions within the long coil domain. Only the talin domain of murine talin has similarity to the Sla2 proteins. (B) Deletion analysis of ScSLA2 domains required for INT1-induced filamentous growth. Illustration of deletion alleles obtained from Reizman and colleagues (not underscored) and Drubin and colleagues (underscored). IIFG, INT1-induced filamentous growth determined using the scoring system described in Table 2.

FIG. 7

CaSLA2 is required for filamentous growth under several different hypha-inducing conditions. CaSLA2/CaSLA2 (YJB1873), CaSLA2/Casla2 (YJB3400), and Casla2/Casla2 (YJB3402) were grown on milk-Tween agar for 5 days (A), in RPMI medium–20% serum for 16 h at 37°C (B), or on YPD agar medium for 13 days (C). Similar results were observed with independently isolated yCA37 and YJB3612 heterozyogous and homozygous Casla2 strains. (C) Parental and two independent Casla2/Casla2 homozygote strains as indicated were photographed after 13 days of growth on YPAD at room temperature.

FIG. 8

Int1p triggers filamentous growth through at least two pathways. Sla2p and Swe1p contribute independently to polarized growth in S. cerevisiae cells expressing INT1. Swe1p may also trigger the morphogenesis checkpoint via Swe1p.