The Molecular Mechanism of White-Opaque Switching in Candida albicans
White-opaque switching is a process by which the opportunistic fungal pathogen Candida albicans reversibly alternates between two growth states, named "white" and "opaque". The white-opaque switch regulates over 400 genes transcriptionally, and cells in each state have different morphology, virulence properties, metabolism, and mating competency. At the time I started my work, there was minimal molecular understanding of the white-opaque switch, and my goal was to identify the genes responsible for generating the opaque state. My work identified a gene, which we named White-Opaque Regulator 1 (WOR1), as the master regulator of the white-opaque switch. This gene is necessary for opaque formation and ectopic expression of WOR1 can drive cells to the opaque state, as verified through an examination of morphology, expression profiling, and behavioral tests. Northern analysis and chromatin immunoprecipitation (ChIP) assays revealed that Wor1 binds its own promoter and upregulates its own expression, forming a self-sustaining positive feedback loop.
Subsequent work sought to understand Wor1 in the context of other transcriptional regulators with known effects on white-opaque switching (CZF1, EFG1, and WOR2). Mutants of CZF1, EFG1, and WOR2 were analyzed in combination with WOR1 to define genetic epistasis relationships. This genetic analysis was combined with ChIP experiments to analyze where Wor1 was associated with chromatin across the entire C. albicans genome. Wor1 was found to associate with the DNA upstream of each of the other regulators. Using the genetic relationships and ChIP data together, we defined a "circuit" of interlocking regulatory feedback loops that are modeled to reinforce Wor1 expression during initiation of the switch to the opaque state.
Unpublished work found that increased osmotic pressure or contact with solid surfaces can increase the frequency of white-to-opaque switching. Ongoing research focuses on purifying Wor1 for use in assays to determine if it can directly bind DNA. Together these studies enriched our understanding of the molecular control of the white-opaque switch.