Principal Investigator Graham Walker
Sinorhizobium melilotiestablishes a symbiosis with alfalfa in which it converts atmospheric nitrogen to ammonia. We are studying the mechanisms by which S. meliloti invades the nodules that it elicits on roots and the functions it requires to establish the chronic intracellular infections of host cells necessary for the symbiosis.
Our extensive genetic analyses of the Sinorhizobium-legume symbiosis led to the discovery that these rhizobia synthesize structurally different, but functionally equivalent, acidic exopolysaccharides that play an essential role in nodule invasion on alfalfa by enabling infection thread initiation and extension. We determined the complete pathway for succinoglycan biosynthesis and the multiple mechanisms for generating the low molecular weight derivatives of succinoglycan and other exopolysaccharides that appear to be the symbiotically active forms. Our recent microarray analysis indicates that appropriate symbiotically active exopolysaccharides act as signals to plant hosts to initiate infection thread formation and that, in the absence of this signal, plants terminate the infection process, perhaps via a defense response.
We discovered that the plant symbiont S. meliloti and the mammalian pathogen Brucella abortus share common molecular functions that are required for the chronic intracellular infections of eukaryotic cells that underlie their respective biological roles. For example, we found that the bacA gene, whose function is required for S. meliloti chronic intracellular infection of plant cells inside the nodule, is present in B. abortus as well, where is required for chronic pathogenesis in BALB/c mice. The limited similarity of BacA to the human X-linked adrenoleukodystrophy protein (a peroxisomal transporter of very long chain fatty acids) led us to the discovery that BacA affects the modification of lipid A by very long chain fatty acids (C28/C30) in both S. meliloti and B. abortus.
We have recently shown that the S. meliloti BluB protein, whose absence results in a symbiotic deficiency, carries out what had been the missing step in vitamin B12 biosynthesis, the synthesis of the “lower ligand” dimethylbenzimidazole (DMB). BluB is a new class of enzyme, a flavin destructase that cannibalizes reduced FMN to make DMB. Crystallographic analysis revealed the relationship between oxidoreductases, which use FMN as a cofactor, and BluB, which uses FMN as a substrate.
Ongoing projects include investigations of the details of BluB function and the molecular nature of the B12 requirement for symbiosis; an examination of a master regulator of symbiosis (CbrA) that is also a principle regulator of the bacterial cell cycle in S. meliloti; characterization of a highly conserved gene found in all bacteria whose product is critical for symbiosis; and the role of non-homologous end-joining (NHEJ) in symbiosis.