Synthesis and Turnover of Basal Level Guauosine Tetraphosphate in Escherichia coZi*

Cultures of Escherichia coli growing exponentially in Trisacetate medium were subjected to nutritional shift-up and the pool size of guanosine 5’-diphosphate-3’-diphosphate (ppGpp) as well as the rates of protein synthesis and net RNA synthesis were determined. In the shift to a rich medium (glucose plus 19 amino acids plus hypoxanthine) the basal level of ppGpp falls immediately with a decay constant suggesting total inhibition of synthesis; there is no ppGpp detectable above background for 30 to 40 min. The net rate of RNA synthesis starts to increase within 1 min of the shift-up and has reached its definitive postshift value well before the pool of ppGpp rises above background level. In a shift-up from Tris-acetate medium to Tris-glucose medium there is a much slower readjustment of the ppGpp pool size without the transient disappearance of the nucleotide. However, in a shift-up to Tris-acetate plus 5 amino acids, a medium which supports the same growth rate as Tris-glucose medium, a dramatic, transient drop in the ppGpp pool level was observed. Relaxed cells exhibit very similar behavior to stringent cells in the same shift-up. Our data argue strongly against an exclusive role for ppGpp in regulating RNA synthesis during nutritional shift-up. The kinetic data of 13H]guanosine uptake into GTP and ppGpp pools were analyzed to determine the rate of ppGpp synthesis. This rate was found to be similar during exponential growth in either Tris-acetate medium or Trisglucose medium. During a shift-down from Tris-glucose to Tris-acetate medium the rate of ppGpp synthesis fell by a factor of 1.5 to 2 and the turnover rate is reduced 6to Sfold, suggesting that the expansion in the ppGpp pool during shift-down cannot be due to derepression of synthesis.