Molecular cloning and characterization of cgt, the Brucella abortus cyclic beta-1,2-glucan transporter gene, and its role in virulence.
The animal pathogen Brucella abortus contains a gene cgt, which complemented Sinorhizobium meliloti nodule development (ndvA) and Agrobacterium tumefaciens chromosomal virulence (chvA) mutants. Complemented strains recovered the presence of anionic cyclic beta-1,2-glucan, motility, tumor induction in A. tumefaciens, and nodule occupancy in S. meliloti, all traits strictly associated with the presence of cyclic beta-1,2-glucan in the periplasm. Nucleotide sequencing revealed that B. abortus cgt contains a 1,797-bp open reading frame coding for a predicted membrane protein of 599 amino acids (65.9 kDa) that is 58.5 and 59.9% identical to S. meliloti NdvA and A. tumefaciens ChvA, respectively. Additionally, B. abortus cgt, like S. meliloti ndvA and A. tumefaciens chvA possesses ATP-binding motifs and the ABC signature domain features of a typical ABC transporter. Characterization of Cgt was carried out by the construction of null mutants in B. abortus 2308 and S19 backgrounds. Both mutants do not transport cyclic beta-1,2-glucan to the periplasm, as shown by the absence of anionic cyclic glucan, and they display reduced virulence in mice and defective intracellular multiplication in HeLa cells. These results suggest that cyclic beta-1,2-glucan must be transported into the periplasmatic space to exert its action as a virulence factor.
Roset
Mara S
MS
Instituto de Investigaciones Biotecnológicas-Instituto Tecnológico de Chascomús, Consejo Nacional de Investigaciones Cientificas y Técnicas, Universidad Nacional de General San Martín, Buenos Aires, Argentina.
Ciocchini
Andrés E
AE
Ugalde
Rodolfo A
RA
Iñón de Iannino
Nora
N
D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002003:Brucella abortus / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000472:pathogenicity*; D002006:Brucellosis / Q000382:microbiology / Q000503:physiopathology; D002460:Cell Line; D003001:Cloning, Molecular; D005260:Female; D005816:Genetic Complementation Test; D005936:Glucans / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009154:Mutation; D017422:Sequence Analysis, DNA; D014774:Virulence; D047071:beta-Glucans
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; C484700:Cgt protein, Brucella abortus; C058770:ChvA protein, Agrobacterium tumefaciens; D005936:Glucans; D047071:beta-Glucans; C056783:ndvA protein, Rhizobium meliloti; C033744:beta-1,2-glucan