Furthermore, signs of premature bacteroid senescence were observed in these nodules. These results suggest that loss of Hfq affects the ability of S. meliloti to survive within the intracellular environment of the host. This phenotype has been reported as a common feature of hfq

mutants of phylogenetically distant pathogenic bacteria [10–12, 15, 21, 22]. Legumes provide invading bacteria with defined and dominant energy sources (i.e. dicarboxylic acids for bacteroids) other than the carbon substrates used for free-living growth in the rhizosphere [47]. Therefore, although the alteration of central metabolic pathways could contribute to different extent to the colonization of developing nodules, Sapitinib order they provide only a partial explanation for the hfq endosymbiotic phenotype. Besides nutrient compounds, invading bacteria has to perceive and respond to a variety of plant signals to successfully colonize legume nodules [27, 28], these include; reactive oxygen species released by the host upon infection [49], peptides likely transported into bacterial cells by the product of the bacA gene to launch bacteroid differentiation [50, 51], the low pH of intracellular compartments

[52] or the microoxic environment demanded by the nitrogenase complex to fix atmospheric nitrogen SC79 cost [38]. Our click here proteomic analysis identified GroEL2, GroEL3, GrpE and IbpA chaperons as deregulated in the 2011-3.4 hfq mutant. Four groESL operons and an additional groEL gene are present in the S. meliloti genome, being the groEL1 required for nodulation and nitrogen-fixation [53, 54]. Thus, it can be speculated that

Hfq-dependent chaperones could help also infective rhizobia to cope with the prolonged stress within the plant host. On the other hand, the transcriptomic profiling revealed that the accumulation of FixK1/FixK2 transcripts is Hfq-dependent. RT-PCR experiments on RNA obtained from cells subjected to more stringent microaerophilic conditions revealed that Hfq-mediated regulation of fixK operates in our assumed aerobic conditions but not in microaerobiosis. In S. meliloti fixK expression is also subjected to indirect autoregulation through the inhibition of the FixL isothipendyl sensor kinase by the FixT protein [55, 56]. Therefore, our findings add another level of complexity to the FixK-dependent regulatory circuit whose biological significance remains to be elucidated. The same RT-PCR experiments showed that Hfq also contributes to the positive regulation of nifA, although transcripts of this gene were still detected in the mutant. Down-regulation of nifA would impact on nitrogenase synthesis, thus explaining the Hfq effects on the onset and probably the efficiency of nitrogen fixation itself in 36%-45% nodules that supported growth and development of the 1021Δhfq-inoculated plants in our assays.