TOS9 Regulates White-Opaque Switching in Candida albicans

Abstract

In Candida albicans, the a1-␣2 complex represses white-opaque switching, as well as mating. Based upon the assumption that the a1-␣2 corepressor complex binds to the gene that regulates white-opaque switching, a chromatin immunoprecipitation-microarray analysis strategy was used to identify 52 genes that bound to the complex. One of these genes, TOS9, exhibited an expression pattern consistent with a "master switch gene." TOS9 was only expressed in opaque cells, and its gene product, Tos9p, localized to the nucleus. Deletion of the gene blocked cells in the white phase, misexpression in the white phase caused stable mass conversion of cells to the opaque state, and misexpression blocked temperature-induced mass conversion from the opaque state to the white state. A model was developed for the regulation of spontaneous switching between the opaque state and the white state that includes stochastic changes of Tos9p levels above and below a threshold that induce changes in the chromatin state of an as-yet-unidentified switching locus. TOS9 has also been referred to as EAP2 and WOR1.White-opaque switching was first observed in a strain of Candida albicans isolated from a fatal bloodstream infection (40). The switch affected the cellular phenotype (1, 41, 42), gene expression (24), and a variety of putative virulence traits (40, 41). It also conferred the capacity to colonize skin (23). An early analysis of clinical strains performed in 1986, however, revealed that only 8% underwent the switch (D. R. Soll, unpublished observation; 44). This observation was enigmatic, since all strains of C. albicans possessed opaque state-specific genes (33). In 2002, Miller and Johnson (31) provided not only an explanation for this enigma but also a role for the whiteopaque transition. They found that while an a/␣ laboratory strain could not switch, MTLa1 and MTL␣2 deletion derivatives of that strain, which were ␣ and a, respectively, could. They demonstrated that switching in the a/␣ strain was repressed by the a1-␣2 complex, the same complex that repressed mating (31). Their results suggested that in order to switch, natural strains, which are predominantly a/␣ (26,30,48), first had to undergo homozygosis to a/a or ␣/␣. Lockhart et al. (30) generalized this observation by demonstrating that while natural a/␣ strains did not undergo white-opaque switching, spontaneously generated MTL-homozygous offspring and natural MTL-homozygous strains did switch. Miller and Johnson (31) further demonstrated that in order to mate, the a and ␣ strains they derived by deleting MTL␣2 and MTLa1, respectively, first had to switch from white to opaque. Lockhart et al. (29) generalized this observation by demonstrating that only natural a/a and ␣/␣ strains that expressed the opaque phenotype could mate. The white-opaque transition, therefore, was an essential and unique step in the C. albicans mating process (3,42,43).In spite of the fundamental role white-opaque switching plays in mating, very little is known about the molecular mechanisms that re...

Stress-Induced Phenotypic Switching inCandida albicans

“…Genes encoded at the mating type-like (MTL) locus provide an additional level of regulation on the whiteopaque switch. The white-opaque transition does not occur in MTLa/␣ cells because the a1/␣2 heterodimer inhibits WOR1 expression, thus locking cells in the white state Tsong et al, 2003;Huang et al, 2006;Srikantha et al, 2006;Zordan et al, 2006). This explains why white-opaque switching is not observed in most clinical isolates of C. albicans; only 3-8% of naturally occurring strains are homozygous at the MTL locus (Lockhart et al, 2002;Odds et al, 2007).…”

“…In the absence of WOR1, cells are unable to form opaques, whereas overexpression of WOR1 forces cells to switch to the opaque state. Stable expression of WOR1 in opaque cells is maintained by positive feedback of Wor1p on its own promoter (Huang et al, 2006;Srikantha et al, 2006;Zordan et al, 2006). Genes encoded at the mating type-like (MTL) locus provide an additional level of regulation on the whiteopaque switch.…”

“…The central factor regulating the white-opaque switch is encoded by the WOR1 gene (also called TOS9/EAP2) (Huang et al, 2006;Srikantha et al, 2006;Zordan et al, 2006). In the absence of WOR1, cells are unable to form opaques, whereas overexpression of WOR1 forces cells to switch to the opaque state.…”

The Same Receptor, G Protein, and Mitogen-activated Protein Kinase Pathway Activate Different Downstream Regulators in the Alternative White and Opaque Pheromone Responses ofCandida albicans

“…Northern and Southern analyses were performed as described previously (Lockhart et al, 2003b;Srikantha et al, 2006). For northern analyses, cells from saturation phase cultures were diluted into fresh medium in the absence or presence of 3 ϫ 10 Ϫ6 M ␣-pheromone, and they were pelleted after 4 h. Total RNA was extracted using the RNeasy Mini kit (QIAGEN, Valencia, CA).…”

Crystal structure of the WOPR-DNA complex and implications for Wor1 function in white-opaque switching of Candida albicans

“…C. albicans can switch between two distinct cell types, white and opaque, which have different properties including cell shape, colonial morphology, metabolic preference, mating ability, gene expression pattern, and host tissue preference [5][6][7][8][9]. White-opaque switching is controlled through expression of a master regulator, Wor1 (whiteopaque switching regulator 1), which is highly upregulated in opaque cells and is required for both the transition to and maintenance of the opaque cell type [10][11][12][13]. Wor1 can genetically interact with the other five key regulators, forming a network of positive and negative feedback loops to control white-opaque switching [14,15].…”