Figure 1.
Genetic organization of RuBisCO 1 (encoded by BRADO1659 and BRADO1660) and RuBisCO 2 (encoded by BRADO2274 and BRADO2275) loci of Bradyrhizobium sp. ORS278 and corresponding synthenic regions of RuBisCO 1 in other rhizobial models.
LysR type regulator are represented in blue, RuBisCO genes in yellow, other CBB cycle genes in purple and carboxysome genes in red. Amino-acid identity level is represented as a percentage for the genes found in synteny.
Figure 2.
Evolutionary relationships of RuBisCO large subunit sequences.
Neighbour joining method was used. Bootstrap support values (10000 replicates with Mega4) are provided as percentage at the corresponding nodes. CbbL1 and CbbL2 proteins from ORS278 strain are highlighted in bold characters. For RuBisCO IA members, the RuBisCO subclasses IAc and IAq is indicated before bacterial name.
Figure 3.
cbbL1 and cbbL2 expression and symbiotic phenotypes of the cbbL1 mutant compared with the WT-strain and the nifK mutant on A. indica.
A. Transcriptional activity of the cbbL1 and cbbL2 putative promoter regions fused to gusA gene revealed on 40 µm nodule sections stained with X-gluc. B. Aerial part and C. root systems of plants inoculated with Bradyrhizobium sp. ORS278 and cbbL1 and nifK derivative strains, white and red asterisk indicate fix minus like and WT-like nodules respectively.
Figure 4.
Identification of CbbL1 (BRADO1659) peptides in bacteroids protein extract by tandem mass spectrometry and database search.
Figure 5.
RuBisCO 1 mutant displays typical traits of Fix- mutant on A. indica but is able to reduce acetylene in vitro.
A. Symbiotic acetylene reduction assays: A. indica plants were inoculated with the indicated Bradyrhizobium sp. ORS278 strain one week after germination. Acetylene reduction activities were measured on individual plants two weeks after inoculation. B. Nodule numbers induced by the indicated strain on A. indica were determined two weeks after infection. C. Acetylene reduction activities were measured during growth on BNM medium supplemented with oxo-glutarate. Error bars represent standard deviations.
Figure 6.
Hypothetical model of ORS278 central metabolism during the symbiotic process.
Upper part of the figure represents the WT and WT-like situation: Plant supplied organic carbon to bacteria which use it via the tricarboxylic acid cycle (TCA) resulting in the production of reduced cofactors (Red Cof). Reduced cofactors are totally oxidized by the respiratory chain at the early stage of nodule formation, then oxygen concentration drops and Calvin cycle drives the electron excess to CO2. Once nitrogenase synthesis has started, electron excess is consumed by the nitrogen fixation process. Lower part of the figure represents the situation with a cbbL1 mutant leading to a Fix–like nodule: electron excess cannot be drive to CO2 therefore, reduced cofactors accumulate and disturbed bacterial metabolism preventing normal nodule development.