The Role Of P15RS And CREPT In Tumor And Wnt Signaling Pathway

RTT103 plays an important role in yeast growth. A double depletion of CTK1 and RTT103 results in synthetic lethality and a double depletion of CTK2 and RTT103 results in synthetic slow growth while a double depletion of REF2 and RTT103 is non-viable in yeast. In this study, we characterized the functions of p15RS and CREPT, which are two human homologues of yeast RTT103 gene.This thesis is composed of two parts: p15RS regulates Wnt/β-catenin signaling pathway and CREPT regulates human tumors growth.p15RS is a p15INK4b related gene, inhibits cell cycle G1/S transition by regulating expression of Cyclin D1, a target gene ofβ-catenin/TCF4/LEF1. Therefore, in the first part, we studied the mechanism that p15RS regulates Wnt/β-catenin signaling pathway. We observed that p15RS interacts withβ-catenin or TCF4, and the interactions are mediated by the RPR domain of p15RS, aa 407-533 ofβ-catenin and aa 1-200 of TCF4. We found that that p15RS colocalizes withβ-catenin or TCF4 in the nucleus by an immunostaining assay. We demonstrated that over-expression of p15RS suppresses canonical Wnt signaling pathway, results in a retarded cell growth and inhibits expression of the Wnt target genes, while depletion of p15RS demonstrated the reverse effects. Furthermore, we found that over-expression of p15RS reduces the interaction ofβ-catenin and TCF4 and inhibits theβ-catenin/TCF4 complex binding to the promoter region of Cyclin D1, but has no effect on the ability of TCF4 binding to DNA.In the second part of the thesis, we explored the biological functions of CREPT. We observed that CREPT is highly expressed in eight human tumors, and the expression of CREPT is correlated with the survival days of patients. We found that over-expression of CREPT results in a rapid cell growth, promotes colony formation, accelerates tumors growth and promotes cell cycle transition from G1 to S phase, while knocking down CREPT demonstrated reversed effects. Further analyses revealed that CREPT up-regulates Cyclin D1 transcription, CREPT binds to the Cyclin D1 promoter and the region before poly A. Interestingly, CREPT regulates Cyclin D1 transcription through promoting formation of a chromatin loop of Cyclin D1 gene: CREPT enhances RNA polymerase II (RNAP II) binding at the region of Cyclin D1 promoter to promote initiation of Cyclin D1 transcription; when RNAP II moves to the region before poly A, the interaction of CREPT with RNAP II prevents RNAP II from moving ahead, and drives it back to the Cyclin D1 promoter thereafter to form a chromatin loop for Cyclin D1 gene. Our findings revealed a mechanism that CREPT regulates cell cycle and promotes tumor growth, and provides basic information for gene diagnosis and gene therapy.