Candida albicans hyphal formation is inhibited by a quorum-sensing molecule, farnesoic acid, which accumulates in the medium as the cells proliferate. We recently showed that Pho81 is essential for the inhibition of hyphal growth by farnesoic acid. Here, we describe a newly identified regulator, Hot1, which increases the expression of PHO81. The binding site of Hot1 in the PHO81 promoter region was identified by DNase I protection assay. The hot1Δ mutant grows extensively as filaments. Furthermore, the inhibition of hyphal formation and the repression of major signaling pathway components in response to farnesoic acid are defective in hot1Δ mutant cells. These data suggest an important role for HOT1 in the inhibition of hyphal development by farnesoic acid in this fungus.