Project description:Background: The overproduction of the main auxin indole-3-acetic acid (IAA) in Sinorhizobium meliloti RD64 leads to beneficial effects both in free-living rhizobium and in host plant during symbiosis. These effects could be related to alteration in gene expression induced by IAA. Preliminary studies of these alterations induced in free-living cells have already started. In this work the transcriptional profiles of free-living S. meliloti wild type 1021 and RD64 strains obtained by RNA-seq analysis were used to better investigate this issue. Results: Transcriptome analysis by next-generation sequencing (RNA-seq) allowed the identification of the genes differentially expressed in RD64 cells, grown in a rich medium under aerobic conditions. Most of the up-regulated genes can be classified into two main categories. The first class comprised the main regulator fixJ and the two intermediate regulators fixK and nifA, which control the expression of genes involved in nitrogen-fixation. For these two latter genes the highest expression levels were observed. Several other genes, known to be FixJ targets, such as those involved in denitrification process and proline metabolism, were also identified. The second class of up-regulated genes comprised the gene coding for the sigma factor RpoH1 and other genes regulated in RpoH1-dependent manner in S. meliloti and involved in stress response. The main category of genes negatively affected can be classified in transport of sugars and other small molecules, chemotaxis and cell envelope functions. Under microaerobic condition, quantitative real-time PCR analysis of genes coding for the two-component regulatory system FixJL and for the regulator NifA showed that for all three genes the induction levels measured in RD64 cells were even higher when compared to aerobic conditions. In particular, for the fixJ gene the expression level increased more than tenfold. The up-regulation of nifA gene was also observed under aerobic conditions both when nitrogen level in the medium was limiting and when it was high. Conclusion: This work provided evidence that the overexpression of the auxin IAA in S. meliloti free-living cells triggers the induction of many genes involved in bacterial response to different abiotic stresses and allows the expression of key genes involved in nitrogen fixation.

Project description:Background: The overproduction of the main auxin indole-3-acetic acid (IAA) in Sinorhizobium meliloti RD64 leads to beneficial effects both in free-living rhizobium and in host plant during symbiosis. These effects could be related to alteration in gene expression induced by IAA. Preliminary studies of these alterations induced in free-living cells have already started. In this work the transcriptional profiles of free-living S. meliloti wild type 1021 and RD64 strains obtained by RNA-seq analysis were used to better investigate this issue. Results: Transcriptome analysis by next-generation sequencing (RNA-seq) allowed the identification of the genes differentially expressed in RD64 cells, grown in a rich medium under aerobic conditions. Most of the up-regulated genes can be classified into two main categories. The first class comprised the main regulator fixJ and the two intermediate regulators fixK and nifA, which control the expression of genes involved in nitrogen-fixation. For these two latter genes the highest expression levels were observed. Several other genes, known to be FixJ targets, such as those involved in denitrification process and proline metabolism, were also identified. The second class of up-regulated genes comprised the gene coding for the sigma factor RpoH1 and other genes regulated in RpoH1-dependent manner in S. meliloti and involved in stress response. The main category of genes negatively affected can be classified in transport of sugars and other small molecules, chemotaxis and cell envelope functions. Under microaerobic condition, quantitative real-time PCR analysis of genes coding for the two-component regulatory system FixJL and for the regulator NifA showed that for all three genes the induction levels measured in RD64 cells were even higher when compared to aerobic conditions. In particular, for the fixJ gene the expression level increased more than tenfold. The up-regulation of nifA gene was also observed under aerobic conditions both when nitrogen level in the medium was limiting and when it was high. Conclusion: This work provided evidence that the overexpression of the auxin IAA in S. meliloti free-living cells triggers the induction of many genes involved in bacterial response to different abiotic stresses and allows the expression of key genes involved in nitrogen fixation. mRNA profiles of S. meliloti wild type 1021 and its derivative IAA-overproducing RD64 strain were generated in triplicate by using HiSeq2000 sequencer

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Comprehensive assessment of the regulons controlled by the FixLJ-FixK2-FixK1 cascade in Bradyrhizobium japonicum

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