CAP1 , an Adenylate Cyclase-Associated Protein Gene, Regulates Bud-Hypha Transitions, Filamentous Growth, and Cyclic AMP Levels and Is Required for Virulence of Candida albicans

Abstract

In response to a wide variety of environmental stimuli, the opportunistic fungal pathogen Candida albicans exits the budding cycle, producing germ tubes and hyphae concomitant with expression of virulence genes, such as that encoding hyphal wall protein 1 (HWP1). Biochemical studies implicate cyclic AMP (cAMP) increases in promoting bud-hypha transitions, but genetic evidence relating genes that control cAMP levels to bud-hypha transitions has not been reported. Adenylate cyclase-associated proteins (CAPs) of nonpathogenic fungi interact with Ras and adenylate cyclase to increase cAMP levels under specific environmental conditions. To initiate studies on the relationship between cAMP signaling and bud-hypha transitions in C. albicans, we identified, cloned, characterized, and disrupted the C. albicans CAP1 gene. C. albicans strains with inactivated CAP1 budded in conditions that led to germ tube formation in isogenic strains with CAP1. The addition of 10 mM cAMP and dibutyryl cAMP promoted bud-hypha transitions and filamentous growth in the cap1/cap1 mutant in liquid and solid media, respectively, showing clearly that cAMP promotes hypha formation in C. albicans. Increases in cytoplasmic cAMP preceding germ tube emergence in strains having CAP1 were markedly diminished in the budding cap1/cap1 mutant. C. albicans strains with deletions of both alleles of CAP1 were avirulent in a mouse model of systemic candidiasis. The avirulence of a germ tube-deficient cap1/cap1 mutant coupled with the role of Cap1 in regulating cAMP levels shows that the Cap1-mediated cAMP signaling pathway is required for bud-hypha transitions, filamentous growth, and the pathogenesis of candidiasis.For many pathogenic fungi, interconversions between morphological growth forms, particularly between yeast growth and filamentous growth, coincide with adaptation to a host environment followed by tissue destruction. Morphological interconversions in fungi are dependent upon signal transduction pathways, including the cyclic AMP (cAMP)-dependent protein kinase A (PKA) pathway (8,10,28,40,46). For the plant pathogens Ustilago maydis and Magnaporthe grisea, cAMP signaling is important for the establishment of filamentous growth in the former and for formation of the infecting appressorium structure of the latter (40, 46). Knowledge about how cAMP signaling mediates morphological interconversion is best understood for Saccharomyces cerevisiae, a budding yeast that produces elongated pseudohyphal cells and forms filamentous colonies in the presence of limiting nitrogen (28, 46). Pseudohyphae exhibit unipolar budding, do not separate, and invade agar (31). Recent experiments involving gene disruption and epistasis analyses have elucidated both upstream and downstream elements of the cAMP-dependent pseudohyphal growth pathway in S. cerevisiae (28,40,46). Adenylate cyclase is activated either through a receptor (Gpr1) that is coupled to a G protein (Gpa2) or by Ras2 (31,41,52,53,58,80). The subsequent activation of PKA then results in activation of the ...

The cAMP phosphodiesterase encoded by CaPDE2 is required for hyphal development in Candida albicans

“…The 1568 bp EcoRI-BglII fragment from pBI-HYHYD (Table 2) was used as probe for EFG1. The 687 bp probe for 18S RNA was generated by PCR using primers recommended by Bahn & Sundstrom, (2001).…”

“…Previous reports on cAMP levels during yeast-to-hypha transition (Egidy et al, 1989(Egidy et al, , 1990Sabie & Gadd, 1992;Bahn & Sundstrom, 2001) are difficult to compare because of differences in strains and indeed experimental conditions. We measured changes in cAMP levels in response to serum with our set of strains (Fig.…”

“…The adenylate cyclase CaCdc35p is essential for hyphal growth and virulence (Rocha et al, 2001). Cap1p and Ras are also required for bud-tohypha transition, cAMP synthesis and virulence (Bahn & Sundstrom, 2001;Feng et al, 1999).…”

Fungal Morphogenesis

“…3) (Cottier and Muhlschlegel 2009). Many of the strong hypha-inducing signals are sensed and integrated by the adenylate cyclase Cyr1, which is indis-pensable for hyphal growth under all conditions (Bahn and Sundstrom 2001;Rocha et al 2001;Hogan and Sundstrom 2009;Zou et al 2009). CO 2 /HCO 3 2 directly stimulates Cyr1p activity by binding to the catalytic domain of Cyr1 (Klengel et al 2005).…”

The Flo8 Transcription Factor Is Essential for Hyphal Development and Virulence inCandida albicans

“…Among them, the cAMP/PKA pathway plays a major role in hyphal development and virulence, because many mutants in the pathway are defective in hyphal growth and show reduced virulence. The adenylate cyclase Cdc35 and its associated protein Cap1 are required for hyphal development under all hyphal-inducing conditions, including serum (Bahn and Sundstrom, 2001;Rocha et al, 2001). The cyclase activity is regulated by two G proteins, Ras1 and Gpa2, in C. albicans (Feng et al, 1999;Sanchez-Martinez and Perez-Martin, 2002;Miwa et al, 2004;Maidan et al, 2005).…”