The heterodimeric HU protein, highly conserved in bacteria and involved in transposition, recombination, DNA repair, etc., shares similarity with histones and HMGs. HU, which binds DNA with low affinity and without sequence specificity, binds strongly and specifically to DNA junctions and DNA containing singlestrand breaks. The fine structure of these specific complexes was studied by footprinting and HU chemically converted into nucleases. The positioning of HUαβ on nicked DNA is asymmetrical and specifically oriented: the β-arm binds the area surrounding the break whereas the α-arm lies on the 3Ј DNA branch. This positioning necessitates a pronounced bend in the DNA at the discontinuous point, which was estimated by circular permutation assay to be 65°. At junctions, HU is similarly asymmetrically positioned in an identical orientation: the junction point plays the role of the discontinuous point in the nicked DNA. The HU binding motif present in both structures is a pair of inclined DNA helices.
SummaryEscherichia coli HU protein is a major component of the bacterial nucleoid. HU stabilizes higher order nucleoprotein complexes and belongs to a family of DNA architectural proteins. Here, we report that HU is required for efficient expression of the sigma S subunit of RNA polymerase. This rpoS-encoded alternative sS factor induces a number of genes implicated in cell survival in stationary phase and in multiple stress resistance. By analysis of rpoS±lacZ fusions and by pulse-chase experiments, we show that the efficiency of rpoS translation is reduced in cells lacking HU, whereas neither rpoS transcription nor protein stability is affected by HU. Gel mobility shift assays show that HU is able to bind specifically an RNA fragment containing the translational initiation region of rpoS mRNA 1000-fold more strongly than double-stranded DNA. Together with the in vivo data, this finding strongly suggests that, by binding to rpoS mRNA, HU directly stimulates rpoS translation. We demonstrate here that HU, an abundant DNAbinding, histone-like protein, is able specifically to recognize an RNA molecule and therefore play a role in post-transcriptional regulation.
HU, a major component of the bacterial nucleoid, shares properties with histones, high mobility group proteins (HMGs), and other eukaryotic proteins. HU, which participates in many major pathways of the bacterial cell, binds without sequence specificity to duplex DNA but recognizes with high affinity DNA repair intermediates. Here we demonstrate that HU binds to doublestranded DNA, double-stranded RNA, and linear DNA-RNA duplexes with a similar low affinity. In contrast to this nonspecific binding to total cellular RNA and to supercoiled DNA, HU specifically recognizes defined structures common to both DNA and RNA. In particular HU binds specifically to nicked or gapped DNA-RNA hybrids and to composite RNA molecules such as DsrA, a small non-coding RNA. HU, which modulates DNA architecture, may play additional key functions in the bacterial machinery via its RNA binding capacity. The simple, straightforward structure of its binding domain with two highly flexible -ribbon arms and an ␣-helical platform is an alternative model for the elaborate binding domains of the eukaryotic proteins that display dual DNA-and RNA-specific binding capacities.The Escherichia coli HU protein is a major component of the bacterial nucleoid (1-3). This small basic histone-like protein that can introduce negative supercoiling into a close circular DNA molecule in the presence of topoisomerase I is highly conserved and found in all bacterial species (4 -7). HU plays a role in DNA replication, recombination, and repair (8 -10). It participates in Mu transposition (11) and regulation of gene transcription (12). HU has been shown to be important for optimal survival of cells in the stationary phase and under various stress conditions (13).HU belongs to the family of architectural nuclear proteins that control DNA topology by introducing bends into doublestranded (ds) 1 DNA and stabilize higher-order nucleoprotein complexes. HU resembles eukaryotic proteins of the high mobility group (HMG) class in its DNA binding properties because it binds dsDNA with low affinity and no sequence specificity. In contrast, it displays high affinity for some altered DNA structures such as junctions, nicks, gaps, forks, and overhangs even under stringent salt conditions (14 -18). The DNA structural motif for HU recognition consists of either two dsDNA modules with propensity to be inclined or one dsDNA module adjacent to a ssDNA binding module (19). X-ray crystallography and NMR studies have established the structure of HU dimer in the absence of DNA (20 -22). The two subunits are intertwined to form a compact ␣-helical hydrophobic core with two extended positively charged -ribbon arms. Our recent studies suggest that HU contacts duplex DNA via the minor groove with its flexible arms, whereas the high affinity binding to its specific binding motif requires an additional contact with the HU body (19). Similar to histones, HU has been shown to bind to poly(U) homopolymer, 2 but the role of this abundant protein in RNA binding was underestimated. Recently we ...
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