Cold shock induces a major ribosomal-associated protein that unwinds double-stranded RNA in Escherichia coli.

Abstract

A 70-kDa protein was specifically induced in Escherichia coli when the culture temperature was shifted from 37 to 15°C. The protein was identified to be the product of the deaD gene (reassigned csdA) encoding a DEAD-box protein. Furthermore, after the shift from 37 to 15°C, CsdA was exclusively localized in the ribosomal fraction and became a major ribosomal-associated protein in cells grown at 15°C. The csdA deletion significantly impaired cell growth and the synthesis of a number of proteins, specifically the derepression of heat-shock proteins, at low temperature. Purified CsdA was found to unwind double-stranded RNA in the absence of ATP. Therefore, the requirement for CsdA in derepression of heat-shock protein synthesis is a cold shock-induced function possibly mediated by destabilization of secondary structures previously identified in the rpoH mRNA.Bacterial adaptation to various environmental stresses has been extensively investigated (reviewed in refs. 1-4). Interestingly, it has been demonstrated that Escherichia coli has an adaptive response not only to high temperature by inducing a group of heat-shock proteins but also to low temperature by inducing a group of cold-shock proteins (5, 6). In contrast to heat-shock proteins, which include protein chaperones required for protein folding and peptidases, cold-shock proteins appear to be involved in various cellular functions such as transcription, translation, and DNA recombination (5, 6).Among the cold-shock proteins of E. coli, CspA has been identified as the major cold-shock protein, which is almost exclusively produced at low temperature at a level of 250,000 molecules per cell (5, 7). The three-dimensional structure of CspA consisting of 69 amino acid residues has been determined, which is composed of five antiparallel (3-sheet structures (8,9). CspA binds to single-stranded DNA (8), and its possible function as an RNA chaperone has been speculated (6). In addition to CspA, E. coli contains a large family of CspA-like proteins consisting of CspB, CspC, CspD, and CspE, among which only CspB is a cold-shock protein (10, 11).In the present paper, we report a newly discovered coldshock protein of 70 kDa, which is also almost exclusively produced upon a temperature shift from 37 to 15°C, similar to the induction of CspA. It was found that this newly identified cold-shock protein is exclusively localized in the ribosomal fraction and became a major ribosomal-associated protein at low temperature. This protein was purified and identified to be the product of the gene that has been known as deaD. This gene had been isolated as a multicopy suppressor for a temperature-sensitive mutation located in the gene encoding ribosomal S2 protein and proposed to encode a putative ATP-dependent RNA helicase based on sequence similarities with other known DEAD-box proteins (12). This protein now is assigned CsdA for cold-shock DEAD-box protein A. We found that this protein has a helix-destabilizing activity. Disruption of the gene resulted in a defect in growth a...

Functional importance of RNA interactions in selection of translation initiation codons

“…In the case of the rpoH-gene-encoded 32 transcription factor, it has been proposed that a highly conserved mRNA secondary structure shielding the initiation codon and DB signal is opened in response to heat shock (Nagai et al, 1991;Nakahigashi et al, 1995). In support of this, the stress-induced ribosomal-associated 'DEAD-box' protein CsdA has been shown to stimulate synthesis of several heat-shock-induced proteins in E. coli, possibly by unwinding the inhibitory structure around the rpoH ribosomebinding site (Jones et al, 1996). Although CsdA has sequence homology to the eukaryotic translation initiation factor eIF-4A (an ATP-dependent RNA helicase), it appears to have the ability to unwind double-stranded RNA in the absence of ATP (Jones et al, 1996).…”

“…In support of this, the stress-induced ribosomal-associated 'DEAD-box' protein CsdA has been shown to stimulate synthesis of several heat-shock-induced proteins in E. coli, possibly by unwinding the inhibitory structure around the rpoH ribosomebinding site (Jones et al, 1996). Although CsdA has sequence homology to the eukaryotic translation initiation factor eIF-4A (an ATP-dependent RNA helicase), it appears to have the ability to unwind double-stranded RNA in the absence of ATP (Jones et al, 1996). Similarly, a conformational switch of the E. coli rpoS ( s ) mRNA structure is likely to improve the accessibility of its ribosome-binding site and stimulate translation of the RNA polymerase cofactor s in response to changing growth conditions (Muffler et al, 1996).…”

RNA remodeling and gene regulation by cold shock proteins

“…The requirement of CsdA at low temperature is more pronounced than that of SrmB and the deletion of csdA gene severely impairs growth at low temperature. 83,84 CsdA has been assigned multiple cellular functions including ribosome biogenesis, translation initiation and degradation of mRNAs.…”

Characterization of the Staphylococcus aureus Heat Shock, Cold Shock, Stringent, and SOS Responses and Their Effects on Log-Phase mRNA Turnover

“…Bacteria cope with conditions of nutrient limitation by eliciting the stringent response, which both reduces the cellular protein synthesis capacity and increases amino acid biosynthesis when substrates for protein synthesis are lacking (42,52). Products of the cold shock response restore translation apparatus function, which is compromised at low temperatures, and resolve low temperature-mediated mRNA secondary structures that would otherwise impede the translation machinery (25,26). At elevated temperatures, cells express heat shock factors that degrade/restructure heat-denatured proteins as well as factors that restore temperature-mediated alterations in chromosome topology (33,61).…”