Effects of Hfq on the conformation and compaction of DNA

Abstract

Hfq is a bacterial pleiotropic regulator that mediates several aspects of nucleic acids metabolism. The protein notably influences translation and turnover of cellular RNAs. Although most previous contributions concentrated on Hfq's interaction with RNA, its association to DNA has also been observed in vitro and in vivo. Here, we focus on DNA-compacting properties of Hfq. Various experimental technologies, including fluorescence microscopy imaging of single DNA molecules confined inside nanofluidic channels, atomic force microscopy and small angle neutron scattering have been used to follow the assembly of Hfq on DNA. Our results show that Hfq forms a nucleoprotein complex, changes the mechanical properties of the double helix and compacts DNA into a condensed form. We propose a compaction mechanism based on protein-mediated bridging of DNA segments. The propensity for bridging is presumably related to multi-arm functionality of the Hfq hexamer, resulting from binding of the C-terminal domains to the duplex. Results are discussed in regard to previous results obtained for H-NS, with important implications for protein binding related gene regulation.

Alpha‐Synuclein Modulates the Physical Properties of DNA

“…The idea was to compare the increase in extension when confinement changes and to use this parameter to determine the physical properties of the protein-bound DNA. [15a] We found the effect on DNA upon aS binding to follow the same trend in the wider channel, that is, the DNA is extended with increasing concentration of aS, but the magnitude of the increase in DNA extension is smaller (Figure 2). This is reflected by an increase in the ratio of the extension at the two confinements as protein is added.…”

“…[24b] In contrast, HU and Hfq interactions induced contraction of the DNA that was proposed to involve protein-mediated bridging of distal DNA segments. [15b,24a] We observed no compaction of DNA by aS at the protein concentrations studied, although AFM show complete coverage of DNA with aS at the highest aS concentration. This may be surprising considering the high positive charge of aS at pH 7.4, but agrees with a similar study on a designed cationic polypeptide.…”

“…Using an analogous approach, thorough single molecule experiments of DNA binding of the bacterial nucleoid proteins HNS, Hfq, and HU have been reported. [15b,22,24] These proteins play pivotal roles in compaction, transcription, and replication of the bacterial genome. As observed here for aS-DNA, H-NS binding to DNA resulted in elongated DNA molecules with increased (doubled) persistence length.…”

“…The degree of stretching is sensitive to the surrounding environment, and to molecules interacting with DNA. Therefore, this method is well suited for studying proteins that cause conformational changes in the DNA, and protein-induced DNA stiffening [14] as well as compaction [15] have been reported…”

Role of Hfq in Genome Evolution: Instability of G-Quadruplex Sequences in E. coli

“…Hfq notably mediates translation efficiency by using stress-related small regulatory RNA (sRNA) and modulates the cellular levels of RNAs, either by changing their stability or through an unsolved transcriptional mechanism [37]. More recent analyses focused attention on the action of Hfq on nucleoid structure [35,[38][39][40][41][42]. These analyses evidenced that Hfq cooperatively binds to DNA through a DNA:protein:DNA bridging mechanism, that it can form filaments on DNA, changing the mechanical properties of the double helix, with important implications for bacterial transcription and replication [35,36].…”