Global regulatory mutations in csrA and rpoS cause severe central carbon stress in Escherichia coli in the presence of acetate

The csrA gene encodes a small RNA-binding protein, which acts as a global r egulator in Escherichia coli and other bacteria (T. Romeo, Mel. Microbiol. 29:1321-1330, 1998). Its key regulatory role in central carbon metabolism, both as an activator of glycolysis and as a potent repressor of glycogen bi osynthesis and gluconeogenesis, prompted us to examine the involvement of c ad in acetate metabolism and the tricarboxylic acid (TCA) cycle; We found t hat growth of csrA rpoS mutant strains was very poor on acetate as a sole c arbon source. Surprisingly, growth also was inhibited specifically by the a ddition of modest amounts of acetate to rich media (e.g., tryptone broth). Cultures grown in the presence of greater than or equal to 25 mM acetate co nsisted substantially of glycogen biosynthesis (glg) mutants, which were no longer inhibited by acetate. Several classes of glg mutations were mapped to known and novel loci. Several hypotheses were examined to provide furthe r insight into the effects of acetate on growth and metabolism in these str ains. We determined that csrA positively regulates acs (acetyl-coenzyme A s ynthetase; Acs) expression and isocitrate lyase activity without affecting key TCA cycle enzymes or phosphotransacetylase. TCA cycle intermediates or pyruvate, but not glucose, galactose, or glycerol, restored growth and prev ented the gig mutations in the presence of acetate. Furthermore, amino acid uptake was inhibited by acetate specifically in the csrA rpoS strain. We c onclude that central carbon flux imbalance, inhibition of amino acid uptake , and a deficiency in acetate metabolism apparently are combined to cause m etabolic stress by depleting the TCA cycle.