RNA helicases: diverse roles in prokaryotic response to abiotic stress - PubMed (original) (raw)
Review
RNA helicases: diverse roles in prokaryotic response to abiotic stress
George W Owttrim. RNA Biol. 2013 Jan.
Abstract
Similar to proteins, RNA molecules must fold into the correct conformation and associate with protein complexes in order to be functional within a cell. RNA helicases rearrange RNA secondary structure and RNA-protein interactions in an ATP-dependent reaction, performing crucial functions in all aspects of RNA metabolism. In prokaryotes, RNA helicase activity is associated with roles in housekeeping functions including RNA turnover, ribosome biogenesis, translation and small RNA metabolism. In addition, RNA helicase expression and/or activity are frequently altered during cellular response to abiotic stress, implying they perform defined roles during cellular adaptation to changes in the growth environment. Specifically, RNA helicases contribute to the formation of cold-adapted ribosomes and RNA degradosomes, implying a role in alleviation of RNA secondary structure stabilization at low temperature. A common emerging theme involves RNA helicases acting as scaffolds for protein-protein interaction and functioning as molecular clamps, holding RNA-protein complexes in specific conformations. This review highlights recent advances in DEAD-box RNA helicase association with cellular response to abiotic stress in prokaryotes.
Keywords: DEAD-box proteins; RNA helicase; abiotic stress; cold-adapted degradosome; cytoskeleton compartmentalization; prokaryotes; ribosome biogenesis.
Figures
Figure 1. DEAD-box RNA helicase structure. The DEAD-box helicase core consists of two RecA-related domains containing a minimum of 12 conserved domains that characterize the family.,,, The sequence of the conserved domains in the E. coli DeaD protein are: Q, PSPIQ; I, GSGKTAAF; Ia, LAPTRELAVQV; Ib, GG; Ic, VGTPGRLLD; II, VLDEADEM; III, FSATM; IV, IIFVRTK; IVa, NGDMNQALR; V, LIATDVA; Va, ARGLDVERISLVVNYD; VI, YVHRIGRTGRAG. The relative order for the domains with RNA-binding or unwinding and ATP binding and/or hydrolysis functions are shown. For example, domain II contains the DEAD motif (Asp-Glu-Ala-Asp). In addition to the helicase core, DEAD-box RNA helicases frequently contain N- and C-terminal extensions that provide RNA and/or protein interaction specificity. The figure is not drawn to scale and has been adapted from Linder and Jankowsky.
Figure 2. RNA helicases and RNA structure remodeling. The majority of RNA helicases are believed to rearrange RNA structure by unwinding dsRNA into ssRNA. Relatively few also catalyze annealing of complementary ssRNA into dsRNA, the annealing and unwinding activities combining to promote RNA strand exchange. These processes require ATP hydrolysis, not for the structure rearrangement but for helicase association with the RNA substrate.,, Unwinding generally occurs over short distances, 1–2 helical turns, however protein cofactors can increase the processivity and also contribute to RNA substrate specificity.
Figure 3. RNA helicases in E. coli ribosome assembly. The relative order of RNA helicase function in ribosome assembly and the sizes of intermediate 50S subunit particles that accumulate in RNA helicase mutants are indicated. RhlE functions early in the assembly process and is proposed to dictate whether assembly will follow an SrmB- or DeaD-specific pathway. Whether the ribosome biosynthetic pathway involves both RNA helicases or if separate SrmB- and DeaD-specific pathways function at high and low temperature, respectively, is not known. SrmB interaction with L4, L24 and the 23S rRNA indicates that it functions before DeaD however there is crosstalk between the two helicases as DeaD overexpression partially rescues the assembly defect in Δ_srmB_ cells (black dotted arrow). SrmB is required to form the L1024 G-ribo wrench pseudoknot, providing the nucleation site for L13 and subsequent events early in ribosome assembly. The role of DbpA in assembly is controversial but is reported to act late in the process, performing a function that occurs in the absence of DbpA, but not in cells expressing both wild type and mutant R331A proteins., DeaD has also been shown to be required for a very late step in 30S assembly involving 16S rRNA.deaD inactivation is partially alleviated by overexpression of cold-induced proteins, RNase R or CspA,, and also RhlE (green dotted arrow), which catalyze RNA unwinding.,, These observations suggest that DeaD functions to remove inhibitory RNA secondary structures whose stability is enhanced at low temperature. The overall objective is to produce stress-condition-specific ribosomes differing in their subunit composition e.g., a cold adapted ribosome. The figure is adapted from Shajani et al.
Figure 4. RNA helicases and stress adapted RNA degradosomes. Subunit composition of the B. subtilis and E. coli degradosome is diagrammed. Under normal and conditions at 37°C, the E. coli degradosome protein subunits are shown with the identified protein-protein interactions indicated by overlapping symbols. The non-catalytic C-terminal domain of RNase E acts as the scaffold for subunit assembly and associates with the membrane via an amphipathic α-helix. RNase E and RhlB independently form helical cytoskeletal structures., RhlB interacts directly with RNase E and PNPase at 37°C. In response to cold stress, RhlB can be exchanged with DeaD (in vitro and in vivo) or RhlE (in vitro) to produce a cold stress degradosome., Stoichiometry of subunits is not known, however a model in which PNPase:RNase E:enolase:RhlB associate in a ratio of 4:3:8:8 binding to the C-terminus of RNase E has been proposed. Similar to E. coli, the B. subtilis and Staphylococcus aureus degradosomes assemble on RNase Y (RNase E equivalent) and are composed of the RNA helicase CshA (RhlB equivalent), enolase and PnpA (PNPase equivalent). In addition the complex contains the RNases J1, J2 and RnpA and the glycolytic enzyme, phosphofructokinase (Pfk)., RNase Y contains a single transmembrane domain, anchoring the complex in the membrane but it is not known if the Bacillus degradosome complex forms helical cytoskeletal structures similar to those observed in E. coli. CshA interacts directly with enolase, Pfk and PnpA however evidence for exchange with other helicases in response to low temperature, although it could not be ruled out, possibly does not occur as CshA levels remain unaltered in response to growth conditions. The diversity of interacting proteins provides the potential for alteration of degradosome composition in response to a range of abiotic stress conditions in B. subtilis.
Figure 5. DEAD box RNA helicases function in the E. coli cold stress response. Association of the five E. coli DEAD-box RNA helicases with cellular pathways associated with cold stress is indicated. DeaD is present at significantly reduced levels at 37°C with expression induced in response to cold stress at 15°C. DeaD helicase activity is associated with a number of cellular pathways. RhlE, SrmB, DeaD and DbpA function in ribosome biogenesis, including 50S and 30S subunit assembly. Whether these helicases function in a linear sequence of reactions associated with 50S subunit assembly or RhlE dictates either an SrmB- or DeaD-specific pathway, as proposed by Jain, remains to be elucidated. At 37°C, RhlE is proposed to direct initial assembly followed by SrmB while at 15°C, RhlE is proposed to direct a DeaD-dependent pathway, possibly associated with formation of a cold-adapted ribosome. Evidence for DeaD functioning at a late stage in 30S assembly has also been presented, aiding association of S1 and thus S2 onto the ribosome. RhlB is an integral subunit of the RNA degradosome at 37°C, catalyzing RNA turnover. The degradosome complex assembles into two interacting cytoskeletal helices, one formed by RNase E and one by RhlB. At 15°C, DeaD and potentially RhlE can functionally replace RhlB, creating a cold-specific degradosome. DeaD also enhances translation at low temperature, potentially by removing RNA secondary structure which prevents translation initiation. However, this effect has been ascribed to reflect DeaD loading of S1 and S2 onto the 30S subunit late in assembly, rather than DeaD unwinding of 5′ UTR secondary structure. Evidence for RNA helicase function in the degradosome catalyzed turnover of sRNA-mRNA complexes, directed to the degradosome by the RNA chaperone Hfq, is circumstantial, although evidence has been presented indicating that RNA helicase activity could be required under specific conditions, for example cold stress. Resch et al. have shown that DeaD is a cofactor required for regulation of rpoS translation by the sRNA, DsrA. At low growth temperature, rpoS translation depends upon DsrA binding which relieves the iss blocking ribosome entry to the rpoS ribosome-binding site. DeaD is proposed to unwind the iss secondary structure prior to Hfq annealing of DsrA to rpoS. It is also possible that DeaD RNA helicase activity extends to a requirement to unfold DsrA and/or Hfq removal once the _rpoS_-DsrA duplex is formed and/or the requirement for an annealing RNA helicase, such as CrhR, to anneal the mRNA-sRNA duplex before interaction with Hfq at low temperature.
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