Cysteine starvation, isoleucyl-tRNAIle, and the regulation of the ilvGEDA operon of Escherichia coli

The involvement of undermodified tRNA in the regulation of the ilvGEDA operon has been investigated using Escherichia coli C6, a relA-, Cys-, Met- mutant. This strain accumulates thionucleotide-deficient or methyl-deficient tRNA when starved for cysteine or methionine, respectively. The levels of threonine deaminase, the ilvA gene product, and transaminase B, the ilvE gene product, were both lower in cysteine-starved cells, as compared with either growing or methionine-starved cultures. When cysteine was added to cysteine-starved cells, growth ensued promptly and both enzyme activities returned to control levels. Treatment of recovering cultures with valine limited growth by isoleucine limitation, but did not cause a derepression of the ilvGEDA operon. Valine treatment of nonstarved or methionine-starved cells led to the expected increase in threonine deaminase and transaminase B activities. Cysteine-starved cells slowly regained the ability to derepress the operon after 3 h of recovery in complete medium. In contrast, the induction of the lac operon was normal in cysteine-starved cultures, even in the presence of valine. The loss of derepressibility of the ilvGEDA operon was correlated with the presence of a kinetically and chromatographically altered tRNAIle in cysteine-starved cells. No changes in tRNAIle were observed after methionine starvation. Using the periodate method, we found that the charging of tRNAIle increased from the normal level of 60 to 80% or greater after starvation for cysteine. Under conditions where the ilvGEDA operon was fully derepressed in nonstarved cells, the charging of tRNAIle fell to 27%. Unexpectedly, nearly identical results were obtained with cysteine-starved cells after an identical derepression test. These results suggest that factors other than the aminoacylation state of tRNAIle may be important in the regulation of this operon. In particular, modifications to tRNA which involve cysteine may be necessary for controlling the expression of the ilvGEDA operon in E. coli.