Positive feedback regulation of E.coli metabolism by HU acetylation mediated by HDAH

Lysine-acetylation (Kac) is a conserved protein post-translational modification (PTM) that plays a key role in regulating protein function in many biological processes such as gene transcription, nutrient metabolism and signal transduction. More and more acylation regulatory enzymes have been discovered in prokaryotes. Here we demonstrate HDAH in Escherichia coli as a potential Kac regulatory enzyme and illustrate the effects of HDAH on bacterial metabolism and growth by mediating Kac of histone-like protein Hu. By using a quantitative proteomic method stable isotope labeling by amino acids in cell culture (SILAC) to screen HDAH-regulated endogenous substrates, we found HDAH can catalyze the removal of HU lysine acetylation marks in vitro and in the cell through a series of biochemical experiments. Next, we used transcriptomic analysis in Escherichia coli and found that downstream genes regulated by HU were mainly related to metabolism. We further demonstrated that the acetylation of HU protein promotes the growth and migration of Escherichia coli by regulating its glucose metabolism. In summary, this study identified a new regulatory enzyme system in bacteria, analyzed the endogenous Kac substrate protein of HDAH, and elucidated the positive feedback regulation molecular mechanism of HU Kac-mediated glycometabolic reprogramming and bacterial proliferation. Overall design: To investigate the effects of HDAH overexpression and HU knockout/mutation on gene expression in E. coli, we constructed BL21 (DE3) strains expressing HDAH using the pET-28a vector, as well as MG1655 strains expressing HU-K22Q and HU-?A for RNA-seq analysis. The samples included E. coli strains overexpressing HDAH, control strains transformed with the empty pET-28a vector, and pET-28a-HU knockout complemented MG1655 strains.