Experimental Studies Of The Relationship Between MFN2 And Osteosarcoma And Its Molecular Mechanism

Objective The tumor supressor role of mitochondrial fusion gene 2(MFN2) have been confirmed in many tumors. However, there are few reports about the mechanism of mfn2 on the development and progression of tumor. We will study the osteosarcoma in this research to reveal the correlation of MFN2 and osteosarcoma. A further demonstration of the potential mechanism between them will be inclued.Methods(1)We collect the pathological specimens of osteosarcoma patients,and then analysis the expression of mfn2 and its effect on the survival of patients with osteosarcoma.(2)A construction of lentiviral which overexpress the mfn2 is needed. We would infect the osteosarcoma cell lines MG63 and U2 OS with the lentiviral and then selected the positive cells with puromcyin to obtain the stable line. The exploration of the overexpression of MFN2 on the proliferation and the apoptosis will be carried out.(3)The purification of FLAG-MFN2 and a mass spectrometry analysis are necessary to detect the interaction protein of MFN2. Whats more, the interaction will be further identified through the immunoprecipitation.(4)The knock out or knock down of MFN2 in the osteosarcoma cell through the CRISPR-cas9 are vital important to our mechnasim analysis.(5)We will study the effect of MFN2 on the development and the progression of osteosarcoma and the potential mechanism, to provide a research foundation for precision treatment.Res?lts(1) The expression of MFN2 is lower in osteosarcoma tissue than the adjacent non tumor tissue both in mRNA and protein level.(2) The stable lines of U2 OS and MG63 which have a high expression of the mfn2 have been screened out through the puromycin selection after the infection of the lentiviral pCDH-FLAG-MFN2.(3) We are the first time identified the PLK1 has an interaction with MFN2 through mass spectrometry analysis. The interaction between MFN2 and PLK1 has been further confirmed through the immunoprecipitation.(4)The stable line cell U2 OS in which the MFN2 is knocked down has been obtained out by the CRISPR-cas9 technique.(5)There has a negative relationship between the expression level of MFN2 and PLK1.Conclusion MFN2 is a novel tumorsupressor for osteosarcoma. And it functions its tumor inhibition roles through the negative regulation of the expression level of PLK1.