OGA Transcriptional Regulates Mitochondrial MT-ATP6 By Affecting NSL3 O-GlcNAc Modification

The degree of cells to adapt to environmental changes is the decisive factor of cell continuous function and survival.This adaptation of cells depends on the accurate coordination,nutrition,metabolism,and environmental factors of the transcriptional program,such as changes in day and night rhythm,which will cause fluctuations in key intermediate metabolites.Cells will perceive the changes in the environment and respond through the changes of these metabolites.The method of response to the environmental response is to use these metabolites to perform a series of modifications to target proteins.These series of modifications that use metabolic products to target proteins are finalized at the level of transcriptional regulation of genes.O-GlcNAc plays a core role in the entire environmental response process,because glucose plays an important role in metabolic response,and in various forms of response,cells' intake of glucose changes.After glucose enters the cell,about 2%?5%enters HBP,and the synthetic OGT substrate UDP-GLc NAc.O-GLc NAc modification of protein is the ultimate manifestation of glucose's role.O-GLc NAc modification is a common post-translational modification(PTM),which is a dynamic,reversible modification of protein-threonic or threonine residues.The signal can be integrated and transmitted in response to different signals within the cell.There are only two enzymes within the cells responsible for O-GlcNAc modifications,respectively,the OGT responsible for adding and the OGA responsible for removal.This study constructed plvx-puro-sh OGA plasmid,and verified knocking down from protein and m RNA levels.Determine OGA knocking after the low feasibility After the MTT,cell scratches,immunofluorescence staining,mitochondrial extraction,RT-q PCR,Western A series of experiments such as blot,verify the impact of the proliferation,invasive transfer capacity of cervical cancer cells after the increase of OGA expression,and changes in mitochondrial related mechanisms involved in this process.This study is divided into the following five parts:1.Successfully constructed plvx-puro-sh OGA plasmid and via RT-q PCR,Western Blot experiments,respectively,of the m RNA and protein levels,respectively.And established the best knockout in 72 hours.2.Using the MTT experiment experiment proves that the expression of cells is promoted after the increase in OGA expression in the 293 T and He La cells.Cell scratches have proved to promote the invasive transfer capacity of the cells after the increase in OGA expression in He La cells.3.Prove with RT-q PCR experiments to increase the expression of OGA in 293 T and He La cells or the expression of NSL3 reduces the transcription of mitochondrial encoded mtDNA.The laboratory pre-data proves that O-GLc NAc modifications can stabilize NSL3,and it is speculated that OGA is regulated by changing the stability of NSL3 to mtDNA transcription.The NSL3 can alleviate the change of mtDNA by knocking down the reply experiment.4.Mitotracker staining,immunofluorescence staining and mitochondrial extraction proof OGA,NSL3,MOF are raised in mitochondrial,suggest NSL3,MOF to regulate mtDNA transcriptional regulation.5.After OGA,O-GlcNAc modification decreased,significantly reduced the recruitment of NSL3 and MOF in mitochondria;after adding OGA inhibitor TMG,the O-GlcNAc modification increased,and the recruitment of NSL3 in mitochondria was significantly reduced.This experiment was concluded,1.Clarifying that O-GLCNAC modification may be caused by NSL3 steady-state changes.2.NSL complex may be used as a sensor in the O-GLCNAC state.The induction external nutritional environment is modified by O-GLCNAC,which in turn causes response and regulate the transcription of mitochondrial oxidized genes.