Event Title
An analysis of Protein that Interact with Xrr1 Protein
Faculty Mentor
Marci Swede
Major/Area of Research
Biomedical Science
Description
Rapamycin, an antifungal and immunosuppressant agent, inhibits cell growth when the FKBP12:rapamycin complex binds to the Tor1 and Tor2 signaling complex, which results in cell cycle arrest. Genetic screens in S. cerevisiae identified mutations in three genes, FPR1 (encodes FKPB12), TOR1 and TOR2 (encoding the Tor proteins), which confer a rapamycin resistant phenotype by allowing cell cycle progression in the presence of the drug. Using the yeast two-hybrid assay, a whole-genome analysis of protein interaction in Saccharomyces cerevisiae yeast revealed a novel gene whose product might interact with the FKB12 (FPR1) protein. This interaction suggests a possible involvement in the rapamycin resistant pathway of this novel gene. Indeed, studies in our laboratory have shown that S. cerevisiae cells carrying a deletion of this novel gene revealed a rapamycin resistance phenotype. Based on this phenotype, the gene was named XRR1 (eXhibits Rapamycin Resistance). The focus of this study is to confirm the putative interaction of the Xrr1 protein and FKBP12 protein in vivo, as suggested by the two-hybrid assay. Our approach was to create a yeast strain that carries epitope tagged versions of the XRR1 and FPR1 genes. This strain was used in co-immunoprecipitation experiments to determine if the two proteins interact. Through this study we were able to create and validate the required strain and were able to detect the two tagged proteins through immunoprecipitation and Western blots. Preliminary co-immunoprecipitation results suggest that the Xrr1 and Fpr1 proteins do interact in S. cerevisiae.
An analysis of Protein that Interact with Xrr1 Protein
Rapamycin, an antifungal and immunosuppressant agent, inhibits cell growth when the FKBP12:rapamycin complex binds to the Tor1 and Tor2 signaling complex, which results in cell cycle arrest. Genetic screens in S. cerevisiae identified mutations in three genes, FPR1 (encodes FKPB12), TOR1 and TOR2 (encoding the Tor proteins), which confer a rapamycin resistant phenotype by allowing cell cycle progression in the presence of the drug. Using the yeast two-hybrid assay, a whole-genome analysis of protein interaction in Saccharomyces cerevisiae yeast revealed a novel gene whose product might interact with the FKB12 (FPR1) protein. This interaction suggests a possible involvement in the rapamycin resistant pathway of this novel gene. Indeed, studies in our laboratory have shown that S. cerevisiae cells carrying a deletion of this novel gene revealed a rapamycin resistance phenotype. Based on this phenotype, the gene was named XRR1 (eXhibits Rapamycin Resistance). The focus of this study is to confirm the putative interaction of the Xrr1 protein and FKBP12 protein in vivo, as suggested by the two-hybrid assay. Our approach was to create a yeast strain that carries epitope tagged versions of the XRR1 and FPR1 genes. This strain was used in co-immunoprecipitation experiments to determine if the two proteins interact. Through this study we were able to create and validate the required strain and were able to detect the two tagged proteins through immunoprecipitation and Western blots. Preliminary co-immunoprecipitation results suggest that the Xrr1 and Fpr1 proteins do interact in S. cerevisiae.