HK1 Inhibits Telomerase Activity To Regulate Cell Senescence Of Lung Adenocarcinoma Via Binding To TERT

Background The incidence of lung cancer in China is increasing year by year,and it is a common clinical malignant tumor.Metabolic reprogramming is one of the important hallmarks of cancer,including many kinds of alterations in glucose metabolism,lipid metabolism,and amino acid metabolism.The changes in glucose metabolism in cancer are mainly characterized by elevated aerobic glycolysis,known as the Warburg effect,which is characterized by enhanced glucose uptake,active glycolysis,and increased lactate metabolites.Warburg effect provides a large amount of energy for the rapid proliferation of cancer cells and provides raw materials for the synthesis of macromolecules such as proteins and lipids,thus promoting cancer growth.Hexokinases,the first step of glucose metabolism,are highly expressed in lung adenocarcinoma and initiate the glycolytic pathway mainly by binding to the outer mitochondrial membrane.Telomerase is mainly responsible for maintaining telomere length in cells.In normally differentiated somatic cells,the gene transcription of telomerase reverse transcriptase(TERT),the main component of telomerase,is inhibited,resulting in inhibition of telomerase activity and a continuous shortening of telomere length with division.When telomeres shorten to a certain limit,the cell undergoes replicative senescence and then dies.In most cancer cells,the abnormal activation of telomerase prolong the telomere length in cancer cells,so that cancer cells break through the limitation of replication and senescence and proliferate indefinitely.TERT is synthesized in the cytoplasm and binds to the molecular chaperones HSP90 and p23 to enter the nucleus via nucleocytoplasmic shuttle,thus participating in the regulation of telomerase assembly and activity.Aerobic glycolysis provides a large amount of energy for the growth of cancer cells and promotes cancer progression.Our preliminary results revealed that altered glycolysis in cancer cells affects telomerase activity,while how glycolysis alters telomerase activity and its specific molecular mechanisms are unclear.Objective The aim of this study was to investigate the mechanisms by which glycolytic metabolism regulates telomerase activity and telomere length in lung adenocarcinoma cells,and to provide new directions for the treatment of clinical lung adenocarcinoma.Methods 1.2DG was used to inhibit glycolysis in A549 cells,and the altered m RNA and protein levels of TERT were detected by RT-q PCR and western blot,and telomerase activity was detected by telomeric repeat amplification protocol(TRAP),and telomere length was detected by terminal restriction fragmentation(TRF).2.The key glycolytic enzyme HK1 bound to TERT was identified by mass spectrometric analysis and verified by co-immunoprecipitation(COIP).After knockdown of HK1 in A549 cells by sh RNA technology,RT-q PCR and western blot were performed to detect the alteration of m RNA and protein levels of TERT.TRAP and TRF were performed to detect the alteration of telomerase activity and telomere length after HK1 was knocked down.Q-FISH was performed to detect the telomere signal intensity at chromosome ends.Results 1.In lung adenocarcinoma cells A549,telomerase activity was increased and telomeres were lengthened after the addition of glycolysis inhibitor 2DG.2.Hexokinase 1,a key enzyme of glycolysis,functions by interacting with TERT.Knockdown of HK1 promoted TERT entry into the nucleus,resulting in increased expression of TERT protein in the nucleus,increased telomerase activity,and telomere lengthening.3.Knockdown of HK1 promoted cellular senescence,which was enhanced by adding telomerase inhibitor BIBR1532 to inhibit telomerase activity.Conclusion 1.Hexokinase 1 downregulates telomerase activity and telomere length in A549 cells by binding to TERT,altering TERT cytosolic localization,and inhibiting TERT entry into the nucleus.2.Knock-down of HK1 promoted senescence and apoptosis in A549 cells,and the level of cell senescence and apoptosis increased after the combination of telomerase inhibitor BIBR1532.