Symbiotic loci of Rhizobium meliloti identified by random TnphoA mutagenesis.
AUTOR(ES)
Long, S
RESUMO
We have developed a system for using TnphoA (TnphoA is Tn5 IS50L::phoA), which generates fusions to alkaline phosphatase (C. Manoil and J. Beckwith, Proc. Natl. Acad. Sci. USA 82:8129-8133, 1985), in Rhizobium meliloti. Active fusions expressing alkaline phosphatase can arise only when this transposon inserts in genes encoding secreted or membrane-spanning proteins. By confining our screening to 1,250 TnphoA-generated mutants of R. meliloti that expressed alkaline phosphatase, we efficiently identified 25 symbiotically defective mutants, all of which formed ineffective (Fix-) nodules on alfalfa. Thirteen of the mutants were unable to synthesize an acidic exopolysaccharide (exo::TnphoA) that is required for nodule invasion. Twelve of the mutations created blocked at later stages of nodule development (fix::TnphoA) and were assigned to nine symbiotic loci. One of these appeared to be a previously undescribed locus located on the pRmeSU47a megaplasmid and to encode a membrane protein. Two others were located on the pRmeSU47b megaplasmid: one was a new locus which was induced by luteolin and encoded a membrane protein, and the other was dctA, the structural gene for dicarboxylic acid transport. The remaining six loci were located on the R. meliloti chromosome. One of these was inducible by luteolin and encoded a membrane protein which determined lipopolysaccharide structure. Three additional chromosomal loci also appeared to encode membrane proteins necessary for symbiosis. The remaining two chromosomal loci encoded periplasmic proteins required for symbiosis.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=211435Documentos Relacionados
- Isolation of orally attenuated Salmonella typhimurium following TnphoA mutagenesis.
- Identification of an iron-regulated virulence determinant in Vibrio cholerae, using TnphoA mutagenesis.
- Regulation of Rhizobium meliloti exo genes in free-living cells and in planta examined by using TnphoA fusions.
- Physical and genetic characterization of symbiotic and auxotrophic mutants of Rhizobium meliloti induced by transposon Tn5 mutagenesis.
- Amino acids in pneumolysin important for hemolytic activity identified by random mutagenesis.