Résumé
Some legume species of the Crotalaria genus are specifically nodulated by methylotrophic bacteria belonging to the Methylobacteriumnodulans species. The feature of this symbiotic bacterium is its ability to oxidize methanol, a property based on the presence of amethanol dehydrogenase enzyme. Despite a good knowledge of this property and its implication in symbiosis, the molecular dialoguebetween M. nodulans and crotalaria podocarpa leading to symbiosis is largely unknown, except the presence of a nodA nodulation gene inthe genome of M. nodulans ORS 2060. To investigate if M. nodulans ORS 2060 produces Nod factors, molecules considered as the majorbacteria-to-plant signals essential for the establishment of rhizobia–legume symbiosis, we identified and sequenced a nodDABCUIJHQcluster from a genomic library of ORS 2060. Phylogenetic analyses of nod genes revealed that M. nodulans ORS 2060 form a branchtogether with Burkholderia tuberum STM678 and a strain of Methylobacterium sp. (4-46) isolated from Lotononis, and distinct from allthe other rhizobia. To analyse the regulation of ORS 2060 nod genes, we constructed a nodA–LacZ promoter fusion to monitor the nodgene expression with various flavonoids. The flavone apigenin was found to be the strongest inducer of nod gene expression inM. nodulans ORS 2060. This latter flavonoid was used to induce ORS 2060, and Nod factors were purified by high-performance liquidchromatography (HPLC) and further characterized by mass spectrometry. One major Nod factor structure was identified as a pentamerof chitin substituted by C18:1 or C16:0 acyl chains on the non-reducing end and 6-O-sulphated on the other end, suggesting a classicsymbiotic dialogue between M. nodulans and C. podocarpa.