Within the latter group several genes with a major role in translation and cellular RNA/protein turnover were differentially regulated in the mutant; SMc01929 coding for RNAseJ, SMc03796 encoding a putative endoribonuclease L-PSP likely involved in mRNAs cleavage, SMa1126, degP4 and degP1 annotated as determinants of different types of proteases, and rplS/rpmA both encoding ribosomal proteins. MLN0128 All these genes except SMa1126 and degP4, were up-regulated in the mutant. As an independent supporting approach to investigate the Hfq function in S. meliloti the proteomic profiling of the wild-type strain
2011 and its hfq MM-102 nmr mutant derivative 2011-3.4 was also determined. Analysis of 24 Coomassie-stained 2D-gels from bacteria grown on TY medium to lag phase (OD600 0.5-0.8) revealed on average 293 spots of which 33 corresponded to individual polypeptides with reliable differential accumulation selleck screening library in the wild-type and mutant strains (see additional file 2: differentially
accumulated proteins in S. meliloti 2011 wild-type and 2011-3.4 insertion mutant derivative). Mass spectrometry (MALDI-TOF) revealed that 28 of these proteins are encoded in chromosomally located genes, 4 in pSymB and only one in the pSymA megaplasmid, thus confirming the major role of Hfq in regulating S. meliloti chromosomal traits (Fig. 2, lower charts). Of these 33 proteins, 21 were over-represented and 12 under-represented in the 2011-3.4 mutant strain. Classification of the differentially expressed proteins according to the S. meliloti 1021 and KEGG databases identified ALOX15 three main functional categories; transport (12 proteins), small molecule metabolism (8) and chaperones and/or stress factors (4) whereas the remaining 9 were catalogued either as involved in translation (i.e. Tig trigger factor and Efp elongation factor P) or as hypothetical conserved proteins with unpredicted function (7) (Fig. 2, lower circle graph). Comparison
of the transcriptomic and proteomic profiles described in this study revealed an overlap of 9 genes identified as differentially expressed in hfq mutants and wild-type strains in both analyses. Their predicted encoded proteins are the periplasmic components of the ABC transporters of myo-inositol (IbpA), fructose (FrcB), α-glucosides (AglE), amino acids (SMc02259), leucine (LivK) and L-amino acids (AapJ and AapP) as well as two enzymes related to myo-inositol catabolism, IolE and IolD. Therefore, regardless the recognized phenotypic differences between the 1021 and 2011 strains both approaches support the general conclusion that Hfq has a major impact in the regulation of transport and metabolism in S. meliloti. Hfq influences central metabolic pathways in S.