Separation And Identification Of HSP27 Interacting Proteins From Human Hepatocellular Carcinoma Cells

ObjectiveHCC is one of the common malignant cancer in the world,and insensitive to chemotherapy and radiotherapy,only 15% can effect a radical cure at present.The rate of recurrence and metastasis is high. The metastasis is one of the most important malignant phenotype and a high degree of orderly process that many complex molecules involved in. Studies have found that heat shock proteins was widely expressed in many human cancers,involved in tumor cell proliferation, differentiation, invasion and metastasis, death and recognition of the immune system. Heat shock protein 27 (HSP27) as an important protein of small heat shock subfamily, in recent years, its relationship with tumor make more and more emphasis,which by through the cell proliferation, differentiation and apoptosis in the regulation of signal transduction involved in a number of the occurrence of tumor invasion and metastasis, the specific mechanism is not yet very clear.But more and more evidence show that apoptosis have the relevance of the existence of tumor metastasis.In our study ,we investigated the HSP27 and its binding protein in order to provide experimental basis for the further explore gene therapy and metastasis of tumors.Methods1.HepG2 .2215 cells were cultured in DMEM nutrient solution/10% calf serum.2.The total proteins from HepG2 .2215 cells were collected by M-PER proteinum lysate.3.HSP27 binding proteins were recovered by antiHSP27 antibody immunoprecipitation with the total proteins from HCC HepG2.2215 cells. After in-solution digested and analyzed by LC-ESI-ITMS/MS , the recovered protein complexes were identified by peptide sequence tags (PST) and database searching.4.Co-immunoprecipitation and Western blot were performed in identity of the expression of HSP90 interacted with HSP27 protein in cells.ResultsThe results show that seven HSP27 binding proteins were first time identified by database searching, which were HSP90,GRP-78 and HSP70 of HSP70 family members, lysyl hydroxylase, Keratin, type I cytoskeletal 9, Keratin, typeâ… cytoskeletal 10, Keratin, typeâ…¡cytoskeletal. The identified seven HSP27-interacting proteins by IP coupled with MS could be classified into three functional categories based on their functions: chaperones,cytoskeleton organization, metabolic enzymes. ConclutionIt can be concluded that HSP27 in HCC in a related proteins involved in the regulation, with some peptides, or small molecule protein interactions, and the participation of control tumor development.