Abstract
BackgroundMany E. coli genes show pH-dependent expression during logarithmic growth in acid (pH 5–6) or in base (pH 8–9). The effect of rapid pH change, however, has rarely been tested. Rapid acid treatment could distinguish between genes responding to external pH, and genes responding to cytoplasmic acidification, which occurs transiently following rapid external acidification. It could reveal previously unknown acid-stress genes whose effects are transient, as well as show which acid-stress genes have a delayed response.ResultsMicroarray hybridization was employed to observe the global gene expression of E. coli K-12 W3110 following rapid acidification of the external medium, from pH 7.6 to pH 5.5. Fluorimetric observation of pH-dependent tetR-YFP showed that rapid external acidification led to a half-unit drop in cytoplasmic pH (from pH 7.6 to pH 6.4) which began to recover within 20 s. Following acid treatment, 630 genes were up-regulated and 586 genes were down-regulated. Up-regulated genes included amino-acid decarboxylases (cadA, adiY, gadA), succinate dehydrogenase (sdhABCD), biofilm-associated genes (bdm, gatAB, and ymgABC), and the Gad, Fur and Rcs regulons. Genes with response patterns consistent with cytoplasmic acid stress were revealed by addition of benzoate, a membrane-permeant acid that permanently depresses cytoplasmic pH without affecting external pH. Several genes (yagU, ygiN, yjeI, and yneI) were up-regulated specifically by external acidification, while other genes (fimB, ygaC, yhcN, yhjX, ymgABC, yodA) presented a benzoate response consistent with cytoplasmic pH stress. Other genes (the nuo operon for NADH dehydrogenase I, and the HslUV protease) showed delayed up-regulation by acid, with expression rising by 10 min following the acid shift.ConclusionTranscriptomic profiling of E. coli K-12 distinguished three different classes of change in gene expression following rapid acid treatment: up-regulation with or without recovery, and delayed response to acid. For eight genes showing acid response and recovery (fimB, ygaC, yhcN, yhjX, ymgABC, yodA), responses to the permeant acid benzoate revealed expression patterns consistent with sensing of cytoplasmic pH. The delayed acid response of nuo genes shows that NADH dehydrogenase I is probably induced as a secondary result of acid-associated metabolism, not as a direct response to cytoplasmic acidification.
Highlights
Many E. coli genes show pH-dependent expression during logarithmic growth in acid or in base
Cytoplasmic pH change The effect of external acidification on the E. coli cytoplasmic pH under our experimental conditions was tested, in order to assess the state of the cytoplasm during our transcriptional profiling
The external pH was acidified by adding HCl (8.5 mM) to cell cultures of E. coli W3110 containing a tetR-YFP plasmid, suspended in media adjusted to pH 7.6
Summary
Many E. coli genes show pH-dependent expression during logarithmic growth in acid (pH 5–6) or in base (pH 8–9). Rapid acid treatment could distinguish between genes responding to external pH, and genes responding to cytoplasmic acidification, which occurs transiently following rapid external acidification. It could reveal previously unknown acid-stress genes whose effects are transient, as well as show which acid-stress genes have a delayed response. Other kinds of genes up-regulated by acid include periplasmic chaperones, inner-membrane and outer-membrane proteins, acetate-stress proteins, and systems utilizing the proton gradient, such as motility and chemotaxis [11]. Base-induced proteins include fermentation pathways generating acidic products, and metabolic complexes that import protons or minimize proton export, such as the F1Fo ATPase and cytochrome d oxidoreductase (cydAB) [11,12]
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