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1.  Candida albicans HWP1 gene expression and host antibody responses in colonization and disease 
Journal of medical microbiology  2006;55(Pt 10):1323-1327.
In vivo expression of the developmentally regulated Candida albicans hyphal wall protein 1 (HWP1) gene was analysed in human subjects who were culture positive for C. albicans and had oral symptoms (n=40) or were asymptomatic (n=29), or had vaginal symptoms (n=40) or were asymptomatic (n=29). HWP1 mRNA was present regardless of symptoms, implicating hyphal and possibly pseudohyphal forms in mucosal carriage as well as disease. As expected, in control subjects without oral symptoms (n=10) and without vaginal symptoms (n=10) who were culture negative in oral and vaginal samples, HWP1 mRNA was not detected. However, exposure to Hwp1 in healthy culture-negative controls, as well as in oral candidiasis and asymptomatic mucosal infections, was shown by the existence of local salivary and systemic adaptive antibody responses to Hwp1. The results are consistent with a role for Hwp1 in gastrointestinal colonization as well as in mucosal symptomatic and asymptomatic infections. Overall, Hwp1 and hyphal growth forms appear to be important factors in benign and invasive interactions of C. albicans with human hosts.
doi:10.1099/jmm.0.46737-0
PMCID: PMC3244616  PMID: 17005778
2.  Genome-Wide Transcriptional Profiling of the Cyclic AMP-Dependent Signaling Pathway during Morphogenic Transitions of Candida albicans▿ †  
Eukaryotic Cell  2007;6(12):2376-2390.
Candida albicans is an opportunistic human fungal pathogen that causes systemic candidiasis as well as superficial mucosal candidiasis. In response to the host environment, C. albicans transitions between yeast and hyphal forms. In particular, hyphal growth is important in facilitating adhesion and invasion of host tissues, concomitant with the expression of various hypha-specific virulence factors. In previous work, we showed that the cyclic AMP (cAMP) signaling pathway plays a crucial role in morphogenic transitions and virulence of C. albicans by studying genes encoding adenylate cyclase-associated protein (CAP1) and high-affinity phosphodiesterase (PDE2) (Y. S. Bahn, J. Staab, and P. Sundstrom, Mol. Microbiol. 50:391-409, 2003; and Y. S. Bahn and P. Sundstrom, J. Bacteriol. 183:3211-3223, 2001). However, little is known about the downstream targets of the cAMP signaling pathway that are responsible for morphological transitions and the expression of virulence factors. Here, microarrays were probed with RNA from strains with hypoactive (cap1/cap1 null mutant), hyperactive (pde2/pde2 null mutant), and wild-type cAMP signaling pathways to provide insight into the molecular mechanisms of virulence that are regulated by cAMP and that are related to the morphogenesis of C. albicans. Genes controlling metabolic specialization, cell wall structure, ergosterol/lipid biosynthesis, and stress responses were modulated by cAMP during hypha formation. Phenotypic traits predicted to be regulated by cAMP from the profiling results correlated with the relative strengths of the mutants when tested for resistance to azoles and subjected to heat shock stress and oxidative/nitrosative stress. The results from this study provide important insights into the role of the cAMP signaling pathway not only in morphogenic transitions of C. albicans but also for adaptation to stress and for survival during host infections.
doi:10.1128/EC.00318-07
PMCID: PMC2168245  PMID: 17951520
3.  Role of Actin Cytoskeletal Dynamics in Activation of the Cyclic AMP Pathway and HWP1 Gene Expression in Candida albicans▿ †  
Eukaryotic Cell  2007;6(10):1824-1840.
Changes in gene expression during reversible bud-hypha transitions of the opportunistic fungal pathogen Candida albicans permit adaptation to environmental conditions that are critical for proliferation in host tissues. Our previous work has shown that the hypha-specific adhesin gene HWP1 is up-regulated by the cyclic AMP (cAMP) signaling pathway. However, little is known about the potential influences of determinants of cell morphology on HWP1 gene expression. We found that blocking hypha formation with cytochalasin A, which destabilizes actin filaments, and with latrunculin A, which sequesters actin monomers, led to a loss of HWP1 gene expression. In contrast, high levels of HWP1 gene expression were observed when the F-actin stabilizer jasplakinolide was used to block hypha formation, suggesting that HWP1 expression could be regulated by actin structures. Mutants defective in formin-mediated nucleation of F-actin were reduced in HWP1 gene expression, providing genetic support for the importance of actin structures. Kinetic experiments with wild-type and actin-deficient cells revealed two distinct phases of HWP1 gene expression, with a slow, actin-independent phase preceding a fast, actin-dependent phase. Low levels of HWP1 gene expression that appeared to be independent of stabilized actin and cAMP signaling were detected using indirect immunofluorescence. A connection between actin structures and the cAMP signaling pathway was shown using hyper- and hypomorphic cAMP mutants, providing a possible mechanism for up-regulation of HWP1 gene expression by stabilized actin. The results reveal a new role for F-actin as a regulatory agent of hypha-specific gene expression at the bud-hypha transition.
doi:10.1128/EC.00188-07
PMCID: PMC2043390  PMID: 17715368
4.  A 368-Base-Pair cis-Acting HWP1 Promoter Region, HCR, of Candida albicans Confers Hypha-Specific Gene Regulation and Binds Architectural Transcription Factors Nhp6 and Gcf1p▿  
Eukaryotic Cell  2007;6(4):693-709.
To elucidate the molecular mechanisms controlling the expression of the hypha-specific adhesin gene HWP1 of Candida albicans, its promoter was dissected and analyzed using a green fluorescent protein reporter gene. A 368-bp region, the HWP1 control region (HCR), was critical for activation under hypha-inducing conditions and conferred developmental regulation to a heterologous ENO1 promoter. A more distal region of the promoter served to amplify the level of promoter activation. Using gel mobility shift assays, a 249-bp subregion of HCR, HCRa, was found to bind at least four proteins from crude extracts of yeasts and hyphae with differing binding patterns dependent on cell morphology. Four proteins with DNA binding activities were identified by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis after separation by anion-exchange and heparin-Sepharose chromatography. One protein with high similarity to Nhp6, an HMG1 family member in Saccharomyces cerevisiae, and another with weak similarity to an HMG-like condensation factor from Physarum polycephalum implicated changes in chromatin structure as a critical process in hypha-specific gene regulation. Proteins with strong homology to histones were also found. These studies are the first to identify proteins that bind to a DNA segment that confers developmental gene regulation in C. albicans and suggest a new model for hypha-specific gene regulation.
doi:10.1128/EC.00341-06
PMCID: PMC1865660  PMID: 17220463
5.  The Adhesin Hwp1 and the First Daughter Cell Localize to the a/a Portion of the Conjugation Bridge during Candida albicans Mating 
Molecular Biology of the Cell  2003;14(12):4920-4930.
The cell wall protein Hwp1 was originally demonstrated to be expressed exclusively in hyphae of Candida albicans and cross-linked to human epithelium by mammalian transglutaminase. Hwp1 is expressed on the walls of hyphae formed by a/α, a/a, and α/α cells. Hence, it is expressed on hyphae independently of mating type. However, Hwp1 is selectively expressed on the wall of conjugation tubes formed by a/a cells, but not α/α cells, in the mating process. This was demonstrated in all possible crosses between four unrelated natural a/a strains and four unrelated α/α strains. In zygotes, Hwp1 is restricted to that portion of the wall of the conjugation bridge contributed by the a/a parent cell. Hwp1 staining further revealed that the first daughter bud that emerges from the conjugation bridge does so from the a/a-contributed portion. Hwp1 expression and localization during the mating process is, therefore, mating type specific, opaque phase specific, and α-pheromone induced. These results indicate that the mating type-specific contributions to the conjugation bridge during the mating process in C. albicans are qualitatively and functionally distinct and that the a/a portion of the bridge, which selectively contains Hwp1, bears the first daughter cell in the mating process.
doi:10.1091/mbc.E03-04-0264
PMCID: PMC284795  PMID: 14565982
6.  Reevaluation of the Role of HWP1 in Systemic Candidiasis by Use of Candida albicans Strains with Selectable Marker URA3 Targeted to the ENO1 Locus  
Infection and Immunity  2002;70(6):3281-3283.
Previous evaluation of HWP1 in systemic candidiasis in CBA/J mice was done with Candida albicans strains with differing genetic locations of URA3 as a result of Ura-blaster mutagenesis. In this study, the presence of HWP1 and the location of URA3 contributed to the severity of murine systemic candidiasis in BALB/c mice.
doi:10.1128/IAI.70.6.3281-3283.2002
PMCID: PMC128023  PMID: 12011025
7.  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 
Journal of Bacteriology  2001;183(10):3211-3223.
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.
doi:10.1128/JB.183.10.3211-3223.2001
PMCID: PMC95223  PMID: 11325951

Results 1-7 (7)