Proc. Natl. Acad. Sci. USA, published online 22 April 2013, doi:10.1073/pnas.1221734110

Candida albicans stochastically alternates between two phenotypic states in a process called 'white-opaque switching'. Nearly 20% of the genome is differentially regulated in the two states, with five transcriptional regulators known to be involved in the switch. To search for additional regulators, Lohse et al. combined RNA-seq transcriptional profiling data of cells in the two different states and ChIP-chip binding data for the master regulator, white-opaque regulator 1 (Wor1), to find genes that were both upregulated in opaque cells and regulated by Wor1. Overexpression of the only gene identified caused switching to the opaque phenotype; given similar behavior to known regulators, the authors named the protein encoded by this gene Wor3. Surprisingly, genetic deletion of Wor3 also promoted the opaque state, altering the expression levels of 125 genes. Analysis of two libraries of 8-mer DNA strands identified a Wor3-specific 5′-ATAACC-3′ DNA-binding sequence. Comparison of these libraries with Wor3 ChIP-chip results highlighted a correlation with the location of Wor1's preferred sequences, suggesting that the two regulators may cooperate. Bioinformatics analysis and structure prediction algorithms identified Wor3 family members but did not indicate homology to any other protein family except through similarity to Wor3's eight CXXC motifs. Phylogenetic analysis suggested that Wor3 evolved and was then lost, given its presence in a subset of fungal genomes only. The discovery of a new participant in fungal state regulation serves as a reminder that surprises lurk in unannotated DNA.