Selective expression of the β‐subunit of nucleoid‐associated protein HU during cold shock in Escherichia coli

Abstract

SummaryExpression of Escherichia coli hupA and hupB, the structural genes encoding the most abundant nucleoid-associated proteins HUa and HUb has been studied during cold shock. This article demonstrates that: (i) transcriptional expression of hupA is blocked following a sudden temperature downshift (from 37∞C to 10∞C), whereas transcription of hupB from the P2 and P3 promoters is maintained at a constitutive level and is activated de novo from the P4 promoter; (ii) all three hupB mRNAs (transcribed from the three natural promoters P2, P3 and P4) become much more stable than the single hupA transcript; and (iii) the hupB transcripts, unlike that of hupA, are efficiently translated in vivo during cold acclimation and can be actively translated in vitro at low temperature. Taken together, the results indicate that during cold shock the expression of the HUb subunit is preferentially stimulated and that of HUa repressed, suggesting that an altered HUa to HUb expression ratio resulting in an increase of HUa/HUb heterodimers and/or (HUb) 2 homodimers may play an important role during cold adaptation. IntroductionHU is the main nucleoid-associated protein in Escherichia coli. This small basic protein, capable of binding both single-and double-stranded DNA displaying a marked preference for cruciform DNA (Pontiggia et al., 1993;Bonnefoy et al., 1994), is present in no less than 30 000 dimers per cell (Wery et al., 2001 and references therein). Upon interaction with DNA, HU forms complexes in which the DNA is characterized by having the same number of superhelical turns as eukaryotic nucleosomes, and can restrain the torsional stress of supercoiled DNA. Indeed, the HUaHUb heterodimers are typical of stationary phase (Claret and Rouvière-Yaniv, 1997). It has been demonstrated that hupA/hupB double-deletion mutants have a lethal phenotype following either cold or heat shock whereas they display no adverse phenotype under normal growth at 37∞C, aside from having a longer lag period preceding exponential growth. These data suggest that the reorganization of the nucleoid structure and/or the changes of gene expression occurring during cellular adaptation to extensive temperature variations may require the participation of HU (Wada et al., 1988;Mizushima et al., 1997). Cold shock response in bacteria encompasses a number of changes in the metabolic state of the cell occurring when the temperature of the culture is suddenly reduced (e.g. in E. coli, from 37∞C to <20∞C). Immediately after the cold stress, the cells enter an acclimation phase during which only a subset of proteins (the 'cold shock proteins'), which can favour the cell adaptation to the new conditions, are synthesized (for reviews, see Jones et al., 1987;1992;Graumann and Marahiel, 1999;Yamanaka, 1999). At the end of the lag phase, normal cellular activities are restored and cell division resumes although with longer generation times.In light of the above-mentioned cold-sensitive phenotype of the hupA/hupB double mutants and of the fact that H-NS, the second most...

Preferential translation of cold-shock mRNAs during cold adaptation

“…The results of these as well as of other experiments (Giangrossi et al 2002; data not shown) clearly demonstrate that the mRNAs used in the present study display only minor stability differences which by no means could explain their several-fold differences in translational activity. Thus, other reasons must be found to explain the observed translational bias; the experiments described below were designed to investigate the nature of such reasons.…”

“…Because the two non-cold-shock mRNAs (hupA and cspD) are translated at 37°C at a level similar to that of the cold-shock hns mRNA, yet to a much lower level than cspA mRNA and are translated very poorly at 15°C (Table 1), we also included in our comparisons three "cold-tolerant" mRNAs originating from the three promoters (P2, P3, and P4) of hupB, the gene encoding the ␤ subunit of HU. The expression of this gene reaches its maximum in cells entering the stationary phase at 37°C (Claret and Rouviere-Yaniv 1997), and although its transcription and translation are stimulated by a cold stress, hupB cannot be regarded as a typical cold-shock gene, because the level of HU␤ does not substantially increase after cold shock (Giangrossi et al 2002). Thus, as indicated by their denomination, the three hupB transcripts are non-cold-shock mRNAs which can be translated to a fairly high level at both low and high temperature.…”

Cold-stress-induced de novo expression of infC and role of IF3 in cold-shock translational bias

“…De novo synthesis of IF3 after cold shock was measured by pulse-chase experiments followed by immunoprecipitation, electrophoretic separation, and quantification by molecular imager essentially as described (Brandi et al 1996;Giangrossi et al 2002).…”

The nucleoid-associated protein HUβ affects global gene expression in Porphyromonas gingivalis

“…Yet, even with a wide variety of functional NAP homologues, E. coli cells lacking HU do exhibit a variety of growth defects. Strains with mutations in both the a and b-subunits have increased sensitivity to UV and ionizing radiation (Boubrik & Rouviere-Yaniv, 1995;Li & Waters, 1998;Miyabe et al, 2000), have a lethal phenotype following cold or heat shock (Giangrossi et al, 2002;Wada et al, 1988), are defective in cell division (Dri et al, 1991) and are altered in outermembrane protein composition (Painbeni et al, 1997). Recently, the HU regulon was determined in E. coli using transcription profiling of strains deficient in the alpha, beta, or both HU subunits (Oberto et al, 2009).…”