Mentor 1
Derek Thomas, Ph.D.
Abstract
Candida albicans is a fungal commensal that resides in the mucosal tissues of approximately 30-70% of the population. Infections in the otherwise healthy individual are rare; however, infection rates can increase for those with certain risk factors such as invasive surgery or compromised immune systems. Systemic candidiasis is a systemic infection with C. albicans and is the fourth most frequent nosocomial infection in the US. The ability to reversibly switch its morphology between yeast cells, pseudohyphae, and true hyphae is integral to the pathogenic potential of C. albicans, playing key roles in biofilm formation and pathogenicity.
This research focuses on Rfg1, a negative regulator of the filamentation process. Rfg1 is believed to function in conjunction with Tup1 to repress genes associated with filamentation; however, the exact mechanism of interaction between the two proteins is unknown. Our current understanding is mostly extrapolated from the model yeast organism Saccharomyces cerevisiae, but there is restricted homology between Rfg1 (65kDa) and the S. cerevisiae equivalent, Rox1 (40kDa). Of the 1.8 kb sequence of Rfg1, 100 bp in the HMG DNA binding domain shares 52% identity with Rox1 and there is little similarity outside the HMG domain. Past research has shown that a S. cerevisiae strain missing both alleles for Rox1 (Δrox1/Δrox1) shows increased sensitivity to caffeine, a purine analog with deleterious effects. Thus, caffeine sensitivity was assayed in a homozygous null Δrfg1/Δrfg1 C. albicans strain, resulting in no visible change in sensitivity. This data supports the functional disparity between CaRfg1 and ScRox despite their shared similarity.
Recent data from our lab has implicated post-translational modification in the control of the interactions between Tup1 and its co-repressors. In silico analysis of Rfg1 with the web-based phosphorylation software NetPhos 2.0 showed multiple amino acid residues in the primary sequence of Rfg1 have the potential to be phosphorylated. Phosphorylation is a form of post-translational modification which can drastically alter the function of a protein. In the case of Rfg1, phosphorylation could have many effects. It could facilitate the dissociation between Rfg1 and Tup1, causing Rfg1 delocalization to the cytoplasm. Conversely, it could localize Rfg1 and increase its affinity for Tup1.
Our future work will begin to analyze variations in post-translational modifications of Rfg1 and interactions with other co-repressors during different filamentation inducing conditions through affinity purification of a polyhistidine - Rfg1 fusion protein, tandem mass spectrometry and protein crosslinking assays. The results from this project and other projects in the lab should significantly further our understanding of the global repression that plays a key role in the regulation of filamentation, which is the most important virulence associated trait in C. albicans.
*This scholar and faculty mentor have requested that only an abstract be published.