Abstract: The ability to switch between different morphological forms is an important feature of Candida albicans, relevant to its pathogenesis. Many conserved positive and negative transcription factors are involved in morphogenetic regulation of the two dimorphic fungi Candida albicans and Saccharomyces cerevisiae. In S. cerevisiae, transcriptional repressor Sfl1 and activator Flo8 function antagonistically in invasive and filamentous growth. We have previously reported Candida albicans Flo8 as a transcription factor essential for hyphal development and virulence in C. albicans. To determine whether a similar negative factor exists in C. albicans, we identified Candida albicans Sfl1 as a functional homolog of the S. cerevisiae sfl1 mutant. Sfl1 is a negative regulator of hyphal development in C. albicans. Deletion of C. albicans SFL1 enhanced filamentous growth and hypha-specific gene expression in several media and growth temperatures. Overexpression of the SFL1 led to a significant reduction of filament formation. Both deletion and overexpression of the SFL1 attenuated virulence of C. albicans in a mouse model. Deleting FLO8 in a sfl1/sfl1 mutant completely blocked hyphal development in various growth conditions examined, suggesting that C. albicans Sfl1 may act as a negative regulator of filamentous growth by antagonizing Flo8 functions. We suggest that, similar to S. cerevisiae, a combination of dual control by activation and repression of Flo8 and Sfl1 may contribute to the fine regulatory network in C. albicans morphogenesis responding to different environmental cues.