The Effect Of PP1α On Wnt/β-catenin Singnal Pathway In Human Osteosarcoma Cell Line

Objective To discovery the role of Protein Phosphatase 1α(PP1α)that plays in Wnt/β-catenin signal pathway in human osteosarcoma cell line (U-2OS).Methods Total RNA was extracted from U-2OS cells and the coding sequence of PP1αgene was amplified by RT-PCR. After its purification, pEGFP-N1-PP1αwas transfected into U-2OS cells for 24h to 48h and its expression was evaluated by Western blot. Dectect the levels of mRNA ofβ-catenin,c-myc,cyclinD1 which includ in Wnt/β-catenin signal pathwayResults The length of specific fragment amplified by PCR PP1αwas 993 bp ,the recombinant plasmids pEGFP-N1-PP1αwas verified by digestion using restriction endonuclease and sequencing results . Western blot revealed that U-2OS cells transfected with recombinant plasmids expressed higher level of PP1αprotein which will enhance the levels of mRNA ofβ-catenin,c-myc,cyclinD1Conclusion The recombinant eukaryotic expression vectors pEGFP-N1-PP1αwas successfully constructed and expressed in U-2OS cells. The level of PP1αprotein in the transfected cells was elevated which will enhance the levels of mRNA ofβ-catenin,c-myc,cyclinD1 that belong to the Wnt/β-catenin singal pathway...