Effect Of Metastasis-associated Gene 1(MTA1)-Mediated Energy Metabolism Reprogramming On Colon Cancer Development And Progression

Objective Normal cells undergo an energy metabolism reprogramming when they transform into tumor cells,which is one of the most common and vital biological characteristics of tumor cells.Metastasis-associated gene 1(MTA1)has been discovered both nucleus and cytoplasm and recently showed a distribution in mitochondria,where cell energy metabolism mainly occurs,suggesting that it may involve the regulation of cell energy metabolism.Therefore,we intend to explore the effects and mechanisms of MTA1-mediated energy metabolism reprogramming during colon cancer development and progression.Methods The datasets including the TCGA pan-cancer transcriptome datasets,the MTA1-knock-out(MTA1-KO)HCT116 cell-derived RNA-seq,and metabolome,and MTA1-binding protein-derived LC/MS identification results were combined to verify if MTA1 is involved in the regulation of metabolic reprogramming in colon cancer.The Seahorse Extracellular Flux Analyzer was utilized to detect the real-time dynamic glycolysis and mitochondrial oxidative phosphorylation(OXPHOS)in tumor cells.The ATP level,the ATP synthase activity and quantity,the glycogen level and other metabolic phenotypes were detected by corresponding kits.The co-localization of MTA1 and mitochondria were determined by immunofluorescence,and the interaction between MTA1 and mitochondrial-associated protein were detected by co-immunoprecipitation.The mitochondria were isolated by differential centrifugation,and the expression of proteins in mitochondria were investigated by immunoblotting and RT-qPCR.Cell proliferation,invasion and colony formation were detected by RTCA,transwell chambers assay and clone formation assay.The expression of DLAT in different datasets and the relationship between DLAT expression and CRC patients' prognosis were verified by bioinformatics analysis using corresponding databases.Results Firstly,the integrated analysis of multi-omics datasets proved that MTA1 involved into the colorectal cancer(CRC)metabolism reprogramming.LC-MS metabolomics analysis also suggested that significantly altered metabolites in MTA1-knock-out cells are enriched in ubiquinone biosynthesis and alanina,aspartate and glutamate metabolism.Secondly,we detected the metabolism-associated phenotype in CRC cell lines.When MTA1 was depleted in HCT116 cells,Extracellular Acidification Rate(ECAR)and Oxygen Consumption Rate(OCR)were significantly decreased,as well as the expression of glycolysis associated genes,the ATP level in cell,the activity and quantity of ATP synthase and the mtDNA copy number,while these phenotypes were rescued when MTA1 was enhanced in HCT116 cells.Furthermore,the 27 MTA1-interacting proteins are mitochondria-related proteins;MTA1 co-localization with mitochondria was seen in HCT116,MCF7,MDA-MB-231 and HeLa cells by IF;MTA1 interacted with ATP5A1 and NDUFS1,which were expressed in mitochondria.Additionally,we screened the malignant phenotypes of HCT116.The colony formation ability and invasion were enhanced in HCT116 cells when MTA1 was overexpressed,and these results were reversed when the cells were treated with Oligomycin A,which is a mitochondrial ATP synthase inhibitor.Finally,the expression of autophagy marker protein LC3II and pAMPK was significantly increased,and mTOR level was reduced in MTA1-KO HCT116 cells after an eight-hour serum starvation,which is inverse in MTA1 over expressed(MTA1-OE)cells,and the difference was abolished when cells were treated with Oligomycin A.Conclusions MTA1,a newly defined mitochondria-localized protein in tumor cells,plays a crucial role in regulating mitochondrial metabolism in colon cancer.It is capable of binding ATP synthase to enhance its activity in CRC cell,which functions to promote the generation of ATP and eventually benefit the growth and invasion of colon cancer cells.