The Roles Of Mechanism Of MTM1 On Zinc Metabolism In Saccharomyces Cerevisiae

Zinc ion plays vital roles in numerous biological processes,including energy generation,neuro-transmission,signal transduction,transportation of nutrients,and maintenance of p H and osmolarity.Defects in the processes of absorption,distribution,utilization,storage and secretion of zinc lead to serious health problems.At present,there are few studies on the interaction between zinc ions and other metal ions relatively,especially single gene regulating zinc and other metal ions.In the process of studying the regulation of nickel ion metabolism by yeast cell MTM1 gene,we found that it simultaneously regulates the metabolic balance of zinc ions.This project will explore the mechanism how MTM1 gene regulates the metabolic balance of zinc ions.In this project,Saccharomyces cerevisiae cell was used as research object and molecular biology and metallomics methods were used to study the regulation mechanism of MTM1 on zinc ion metabolism.The MTM1 gene was knocked out and the growth difference between the knockout strain and the wild-type yeast was compared under excess zinc conditions and the mechanism was further elucidated.Cell growth experiment showed that the MTM1 knockout strain grow significantly better than the wild type on SC medium supplemented with excess zinc ions,accordingly,overexpression of the MTM1 gene showed sensitivity to zinc.The results of ICP-MS showed that the zinc ion content of the mtm1?mutant strain was dramatically lower than that of the wild type strain.RT-PCR and ?-galactosidase gene reporter results indicated that the transcription level of ZRC1 in the mtm1? mutant was dramatically higher than that in the wild-type strain.The zrc1?mtm1? double knockout strain was constructed by homologous recombination.The growth assay suggested that the double knockout strain was more sensitive to excess zinc than the ZRC1 single knockout strain.The activity of superoxide dismutase 2(SOD2)in mtm1? was severely lower than that of wild type and excess zinc ion decreased the activity of SOD2.Western blotting demonstratedthat zinc ions induce the expression of MTM1 protein.Zinc ions shows no significant effect on the localization of MTM1 protein by green fluorescent protein assay.The results of this study demonstrate that the MTM1 gene regulates the metabolism of zinc ions in eukaryotes by regulating the expression level of ZRC1 gene,which leads to enhancing the efficiency of zinc ion transport from cytoplasm to vacuole and reducing the accumulation of excess zinc ions in the cytoplasm.Finally,the toxic effect of zinc ions on cells is decreased contributing the mtm1? strain exhibits high resistance to zinc ions.The clarification of this mechanism provides new insights and avenues for the study of zinc-related diseases.