Functional characterization of Rbp1p interacting protein, Nrp1p, in Saccharomyces cerevisiae

• Main Authors: Ya-Hui Lin, 林亞慧
• Other Authors: 李芳仁
• Format: Others
• Language: en_US
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/57790006362753431621

碩士 === 國立臺灣大學 === 分子醫學研究所 === 92 === In eukaryotic cells, mRNAs undergo sequential processing reactions, including capping, splicing, and polyadenylation. RNA-binding proteins play key roles in these post-transcriptional regulations of gene expression. Our lab has recently demonstrated that Rbp1p can regulate porin mRNA stability through binding to 3’-UTR of porin mRNA. Here, we show that Nrp1p interacts with Rbp1p in two-hybrid interaction analysis. NRP1 encodes a 720 amino-acid protein, Nrp1p, which contains a RRM, two Zn finger motifs, and an ASN-rich region. By co-immunoprecipitation analysis, we showed that Nrp1p could pull down Rbp1p, and vice versa. Tandem affinity purification also showed that endogenous Nrp1p and Rbp1p appeared in the same protein complex. RNA analysis demonstrated that level of porin mRNA in nrp1 mutant strain was lower than that of wild-type strain, and immuno-fluorescence staining shows that porin expression level was enhanced when overexpressed Nrp1p, suggesting that Nrp1p was involved in porin mRNA metabolism. Moreover, subcellular localization analysis showed that Nrp1p, like Rbp1p, is localized to perinuclear region and partially to mitochondria. In conclusion, our results demonstrate that Nrp1p is an Rbp1p interacting protein and is involved in porin mRNA metabolism cooperatively.