The interaction of the Myb1 and CypA-1 proteins in Trichomonas vaginalis

• Main Authors: Ya-Ting Wang, 王雅婷
• Other Authors: Shiou-Jeng Ong
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/85988306781013805744

碩士 === 國立臺灣大學 === 微生物學研究所 === 95 === Expression of the adhesion protein ap65-1 gene in the human pathogen, Trichomonas vaginalis, is critically regulated by iron, which was shown to enhance the nuclear translocation of Myb1 that may inhibit iron-activated transcription of the ap65-1 gene. In this study, bacterioMatch Two-Hybrid System (Stratagene) was applied to identify potential Myb1 interaction proteins. One of Myb1 interacting partner proteins was identified, and named CypA-1. CypA-1 is a cyclophilin-like protein, a family of highly conserved ubiquitous proteins in all the organisms. Specific interaction between CypA-1 and Myb1 was confirmed by the GST-pull down assay. CypA-1 mRNA level increased with iron treatment. Recombinant CypA-1 was assayed for cis-trans peptidyl-prolyl isomerase (PPIase) activity, and gave a value of kcat/Km 103 M-1S-1. R63A retained 60 % of the catalytic efficiency (kcat/Km) of wild-type recombinant CypA-1. Based on the immunofluorescence staining and Western blot assay, both CypA-1 and R63A were expressed in the cytoplasm. Expression of R63A inhibited the nuclear translocation of Myb1. The region of Myb1 interacting with CypA-1 was found to be E104-I112. YGP105-107AAA also interacted with CypA-1 with decreased activity as reveal by the GST-pull down assay. Based on the immunofluorescence staining, Myb1 expressed in the nucleus, P107A was accumulated close to the nuclear membrane, but YGP105-107AAA was evenly distributed in the cytoplasm. Results indicate the participation by Y105-P107 in the translocation of Myb1. Myb1 was either expressed as accumulated close to nuclear membrane, or in the cytoplasm by cyclosporin A treatment. These results provide a new aspect for investigation on the role of the Myb1 protein in transcription regulation of Trichomonas vaginalis.