DRP1 Regulates The Glucose Metabolism Pathway Through BMAL1 To Promote The Proliferation And Invasion On Breast Cancer Cells

At present,breast cancer has become the most threatening malignant tumor to women's life and health.What's more,the incidence rate of breast cancer is constantly on the rise.The invasion and metastasis of breast cancer often lead to poor prognosis and low survival rate of patients.Previous studies have shown that DRP1 is highly expressed in breast cancer,and abnormally DRP1 expression is also related to poor prognosis of breast cancer.As a dynamin-related protein,DRP1 has been shown to play an important role in mitochondrial fission and peroxisome fission.As the core transcription factor of circadian clock,BMAL1 is essential for glucose metabolism and cell survival.the disorder of the circadian clock cycle will cause abnormal expression of intracellular proteins,which will lead to diseases.Although some literatures have shown that the increases of mitochondrial fission can promote the development of cancer cells,the role of DRP1 in the occurrence and development of breast cancer is less understood.This study mainly finds how DRP1 affects circadian clock-related protein expression,and the influences of this process on the regulation mechanism of DRP1 on the occurrence and development of breast cancer.Firstly,we teste the protein levels of BMAL1 and DRP1 in several breast cancer cell lines(MCF-7,T47 D,ZR-75-30,and MDA-MB-231),and find that the expressions of them are obviously positively correlated,and both BMAL1 and DRP1 are highly expressed in highly aggressive cell lines.The results proved that DRP1 could promote the invasion and metastasis of breast cancer cells using Tumour Invasion Assay.And DRP1 could also increased the proliferation of breast cancer cells using MTT and Clone conformation.Secondly,we teste endogenous BMAL1 in ZR-75-30 cell line with overexpression of DRP1,through Real-time quantitative PCR and westernbloting.It was proved that DRP1 could increase the expression of BMAL1 at the mRNA level and protein level.DRP1 up-regulate the activity of BMAL1-Luc using Luciferase reporter gene assay.The results show overexpression of DRP1 up-regulate the expression of Circadian Proteins at mRNA and protein level in ZR-75-30 cell line,such as CLOCK and target proteins CRY1/2 and PER1/2.Thirdly,to tested function of BMAL1,we interfering BMAL1 expression in overexpression of DRP1 cell line.The results show that the function of DRP1 depend on BMAL1 expression in invasion and metastasis of breast cancer cell lines.Finally,we test the influence of DRP1 in Mitochondrial function.The results show that the mitochondrial fission protein DRP1 increased the protein level of glycolysis HK2 which is the glucose metabolism gene by activating BMAL1 and further promoted the development of breast cancer,using glucose measurement kit and ATP measurement kit.The results verified that the mitochondrial fission protein DRP1 promote the proliferation and invasion of breast cancer,mainly by up-regulating the mRNA and protein levels of BMAL1,further activating the expression of the key glycometabolism proteins HK2 ?PFK1 and LDAH1 thus up-regulating the glycolytic metabolism activity.This study find relationship between mitochondrial division and biological rhythm,and provide further theoretical support for the occurrence and development of breast cancer.