# Riboregulation in Nitrogen-Fixing Endosymbiotic Bacteria
## 6 Proteins Assisting sRNA Activity
### The Rhizobial RNA Claperone Hfq  
The host factor for the replication of the Q\(\beta\) bacteriophage or Hfq is encoded by 55% of the bacterial genomes sequenced so far [111]. The function of this protein as RNA chaperone extends far beyond its primary role in intracellular RNA phage multiplication, to assist regulation by sRNAs (preferentially _trans_-sRNAs) and stabilize diverse RNA species including mRNAs and tRNAs [14]. These RNA-binding features place Hfq at a node of bacterial post-transcriptional regulatory networks. All \(\alpha\)-rhizobial genomes host an Hfq coding gene that appears to be strongly expressed and autoregulated at the translational level [112; 113; 114]. The rhizobial Hfq polypeptide has a C-terminal region shorter than that of its enterobacterial counterparts, but retains the core RNA-binding domains of canonical Hfqs [14; 115]. This predicts similar quaternary structural arrangements of rhizobial and enterobacterial Hfq proteins into ring-shaped homohexamers that expose two distinct surfaces for RNA binding. Accordingly, _E. coli_ and _S. meliloti_ Hfq homologs have been shown to be functionally interchangeable [112]. Like in enterobacteria, _Hfq_ knock-out causes severe pleiotropic phenotypes in _S. meliloti_ that include altered growth, increased sensitivity to abiotic stress, changes in the profile of QS signals, reduced competitiveness for modulation, compromised survival of bacteroids inside nodule cells and defects in N fixation [80; 114; 115; 116; 117]. Late symbiotic deficiencies of these mutants are most likely related to the positive contribution of Hfq to the post-transcriptional control of genes encoding the master regulators of N fixation NifA and FixK in diverse free-living and symbiotic \(\alpha\)-proteobacterial diazotrophs by mechanisms that are still poorly understood [113; 114; 115; 117; 118]. Nonetheless, this phenotypic pleiotropy can be further explained by the profound Hfq-dependent changes observed in the rhizobial transcriptome and proteome upon free-living growth, as recurrently revealed by several independent studies on _S. meliloti_[119; 115; 116; 80]. One remarkable observation common to all these reports is the pervasive misregulation of genes encoding ABC transport proteins in bacteria lacking Hfq, with overall upregulation of those presumably involved in uptake of aa and other N sources, such as the broad specificity Aap and Bra systems [120; 70]. Although probably involving both common and host-specific regulatory mechanisms, the central role of Hfq in the chronic infection of eukaryotic organisms has been also reported in plant and mammal pathogens within the \(\alpha\)-subgroup of proteobacteria such as _A. tumefaciens_ or _Brucella_ species [121; 122; 61]. Regulation of at least part of the Hfq-dependent mRNAs is expected to involve its cognate partner sRNAs. In _S. meliloti_, genome-wide profiling of RNA species co-immunoprecipitation with a functional FLAG epitope-tagged Hfq variant (CoIP-RNA) uncovered massive binding of the protein to mRNAs [32; 123]. Specifically, nearly 20% of predicted _S. meliloti_ mRNAs were enriched in Hfq CoIP-RNA, most of which functionally related to major symbiotic and stress-response pathways. Many of these mRNAs, such as those encoding NifA, FixK or stress-related alternative RNA polymerase \(\sigma\) factors of extracytoplasmic function (e.g., RpoE2) are positively regulated by Hfq by a mechanism that probably involves protection of the transcripts from degradation by cellular ribonucleases in a _trans_-acting sRNA-independent manner [103]. Conversely, mRNAs encoding aa transporters that are predicted or experimentally confirmed targets of the functionally characterized _trans_-sRNAs AbcR1 and AbcR2 are particularly abundant among the Hfq-binding transcripts. In these cases, Hfq most likely acts promoting both _trans_-sRNA stability and base-pairing to mRNA targets to boost post-transcriptional silencing [103]. Nonetheless, only 14% of _trans_-sRNAs and 2% of asRNAs currently annotated in the _S.