Studies on the Mechanism of Morphogenetic Transition Signaling in Candida albicans by Farnesoicacid, a Quorum-sensingMolecule

Abstract

The ability of Candida albicans to switch between yeast and filamentous forms in response to environmental conditions has been postulated to contribute to the virulence of this organism. C. albicans hyphal formation is inhibited by a quorum-sensing molecule, farnesoic acid, which accumulates in the medium as the cells proliferate. Many signaling pathways and regulators involved in morphogenetic transition have been identified from intensive investigations, but the molecular networks of farnesoic acid remain poorly understood. It was recently demonstrated that Pho81, a cyclin-dependent protein kinase inhibitor, is for the inhibition of hyphal formation by farnesoic acid. The pho81 mutant grew exclusively as filaments under the conditions tested. In this study, it was described a newly identified regulator, Hot1, which increases the expression of PHO81. By screening a cDNA library with a yeast one-hybrid assay, the C. albicans gene HOT1 was identified, which encodes a protein homologous to the Saccharomyces cerevisiae Hot1, an osmostress transcription factor. Sequencing and conceptual translation of HOT1 full-length cDNA revealed an open reading frame (ORF) consisting of 607 amino acid with 19% identity to Hot1 from S. cerevisiae over its entire length. The Gcr1_C domain (residues 502–579) was conserved, and the amino acid sequence showed 43% homology with ScHot1. The binding site of Hot1 in the PHO81 promoter region was investigated by electrophoretic mobility shift assay (EMSA) and DNase I protection assays. The region extending from −479 to −449 relative to the binding site of HOT1 was identified. To investigate the biological role of HOT1 in inhibition of hyphal development by farnesoic acid in C. albicans, the hot1 mutant strain cells were constructed, and the morphological characterization were compared to wild-type cells. The hot1 mutant strain grew extensively as filaments and was insensitive to farnesoic acid, but not in the wild-type cells. The hot1 mutant cells expressing CaHOT1 under control of the ADH1 promoters restored a wild-type phenotype. From these results, it was concluded that CaHOT1 had a role in supressing filamentous growth and was a novel molecule for inhibition of the hyphal development by farnesoic acid. Two-hybrid experiments demonstrated that Hot1 interacts with Hog1 mitogen-activated protein kinase (MAPK). Analysis of the expression levels of major signaling pathway components by reverse transcription-polymerase chain reaction (RT-PCR) indicated that farnesoic acid inhibits hyphal formation in C. albicans through the coordination of HOG MAPK pathway. These findings suggest that Hot1 is the regulator of PHO81 transcription and a significant component of filamentation by farnesoic acid in C. albicans.