Global effects of homocysteine on transcription in Escherichia coli: induction of the gene for the major cold-shock protein, CspA.

Homocysteine (Hcy) is a thiol-containing amino acid that is considered to be medically important because it is linked to the development of several life-threatening diseases in humans, including cardiovascular disease and stroke. It inhibits the growth of Escherichia coli when supplied in the growth medium. Growth inhibition is believed to arise as a result of partial starvation for isoleucine, which occurs because Hcy perturbs the biosynthesis of this amino acid. This study attempted to further elucidate the inhibitory mode of action of Hcy by examining the impact of exogenously supplied Hcy on the transcriptome. Using gene macroarrays the transcript levels corresponding to 68 genes were found to be reproducibly altered in the presence of 0.5 mM Hcy. Of these genes, the biggest functional groups affected were those involved in translation (25 genes) and in amino acid metabolism (19 genes). Genes involved in protection against oxidative stress were repressed in Hcy-treated cells and this correlated with a decrease in catalase activity. The gene showing the strongest induction by Hcy was cspA, which encodes the major cold-shock protein CspA. RT-PCR and reporter fusion experiments confirmed that cspA was induced by Hcy. Induction of cspA by Hcy was not caused by nutritional upshift, a stimulus known to induce CspA expression, nor was it dependent on the presence of a functional CspA protein. The induction of cspA by Hcy was suppressed when isoleucine was included in the growth medium. These data suggest that the induction of CspA expression in the presence of Hcy occurs because of a limitation for isoleucine. The possibility that Hcy-induced cspA expression is triggered by translational stalling that occurs when the cells are limited for isoleucine is discussed.