Roles of KLF4 and AMPK in the inhibition of glycolysis by pulsatile shear stress in endothelial cells

Significance This work identifies mechanotransduction mechanisms by which blood flow regulates glycolysis in vascular endothelium. We demonstrate that atheroprotective flow pattern decreases glycolysis, an energy-demanding metabolic process, in endothelium in vitro and in vivo. GCKR, an inhibitor of glycolytic flux, is up-regulated by atheroprotective flow, in contrast to the down-regulation of other glycolysis genes. As a pioneer transcription factor induced by atheroprotective flow, KLF4 epigenetically remodels the GCKR promoter and thus transactivates GCKR. At the posttranslational level, atheroprotective flow–activated AMPK phosphorylates GCKR and hence increases GCKR binding to hexokinase, the key enzyme in glycolysis. The translational significance of these findings builds on the identification of these atheroprotective mechanisms in an animal model with a high level of voluntary wheel running. Vascular endothelial cells (ECs) sense and respond to hemodynamic forces such as pulsatile shear stress (PS) and oscillatory shear stress (OS). Among the metabolic pathways, glycolysis is differentially regulated by atheroprone OS and atheroprotective PS. Studying the molecular mechanisms by which PS suppresses glycolytic flux at the epigenetic, transcriptomic, and kinomic levels, we have demonstrated that glucokinase regulatory protein (GCKR) was markedly induced by PS in vitro and in vivo, although PS down-regulates other glycolysis enzymes such as hexokinase (HK1). Using next-generation sequencing data, we identified the binding of PS-induced Krüppel-like factor 4 (KLF4), which functions as a pioneer transcription factor, binding to the GCKR promoter to change the chromatin structure for transactivation of GCKR. At the posttranslational level, PS-activated AMP-activated protein kinase (AMPK) phosphorylates GCKR at Ser-481, thereby enhancing the interaction between GCKR and HK1 in ECs. In vivo, the level of phosphorylated GCKR Ser-481 and the interaction between GCKR and HK1 were increased in the thoracic aorta of wild-type AMPKα2+/+ mice in comparison with littermates with EC ablation of AMPKα2 (AMPKα2−/−). In addition, the level of GCKR was elevated in the aortas of mice with a high level of voluntary wheel running. The underlying mechanisms for the PS induction of GCKR involve regulation at the epigenetic level by KLF4 and at the posttranslational level by AMPK.