Identification and Characterization of a Gene on Rhizobium meliloti pSyma, syrB, That Negatively Affects syrM Expression

The Rhizobium meliloti SyrM protein activates transcription of nodD3 and syrA. Regulation of syrM is complex and may involve as yet undiscovered genes. Here we report the isolation of insertion mutants showing increased expression of a syrM-gusA gene fusion. Characterization of one mutant strain, designated SYR-B, revealed a mutation consisting of a transposon insertion linked to a large deletion. The corresponding wild-type DNA was cloned as a 5.3-kb BamHI fragment. Genetic and physical analysis of this DNA demonstrated that an open reading frame (ORF) near one end of the fragment, encoding the 16.5-kDa SyrB protein, is responsible for the repression of syrM activity. Results of complementation experiments with the 5.3-kb BamHI DNA led us to hypothesize that other genes within this DNA fragment interfere with the expression or activity of SyrB. Our analysis showed that the region upstream of syrB contains three ORFs. One ORF is similar to the Ros repressor of Agrobacterium tumefaciens and the MucR repressor of R. meliloti.

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Extended Region of Nodulation Genes in Rhizobium meliloti 1021. I. Phenotypes of Tn5 Insertion Mutants

Genetics ◽

10.1093/genetics/117.2.181 ◽

1987 ◽

Vol 117 (2) ◽

pp. 181-189

Author(s):

Jean A Swanson ◽

Janice K Tu ◽

Joy Ogawa ◽

Raghu Sanga ◽

Robert F Fisher ◽

...

Keyword(s):

Rhizobium Meliloti ◽

Large Deletion ◽

Open Reading Frames ◽

Gene Region ◽

Wild Type ◽

Transposon Insertion ◽

Host Range Determination ◽

Range Determination ◽

Mutant Phenotypes ◽

Insertion Mutants

ABSTRACT Rhizobium meliloti Nod- mutant WL131, a derivative of wild-type strain 102F51, was complemented by a clone bank of wild-type R. meliloti 1021 DNA, and clone pRmJT5 was recovered. Transfer of pRmJT5 conferred alfalfa nodulation on other Rhizobium species, indicating a role in host range determination for pRmJT5. Mutagenesis of pRmJT5 revealed several segments in which transposon insertion causes delay in nodulation, and/or marked reduction of the number of nodules formed on host alfalfa plants. The set of mutants indicated five regions in which nod genes are located; one mutant, nod-216, is located in a region not previously reported to encode a nodulation gene. Other mutant phenotypes correlated with the positions of open reading frames for nodH, nodF and nodE, and with a 2.2-kb Eco RI fragment. A mutant in nodG had no altered phenotype in this strain. One nodulation mutant was shown to be a large deletion of the common nod gene region. We present a discussion comparing the various studies made on this extended nod gene region.

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Identification and Characterization of Large Plasmids in Rhizobium meliloti using Agarose Gel Electrophoresis

Journal of General Microbiology ◽

10.1099/00221287-113-2-229 ◽

1979 ◽

Vol 113 (2) ◽

pp. 229-242 ◽

Cited By ~ 310

Author(s):

F. CASSE ◽

C. BOUCHER ◽

J. S. JULLIOT ◽

M. MICHEL ◽

J. DENARIE

Keyword(s):

Gel Electrophoresis ◽

Rhizobium Meliloti ◽

Agarose Gel ◽

Agarose Gel Electrophoresis ◽

Large Plasmids ◽

Identification And Characterization

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Identification and characterization of the first large deletion of theMYH9gene associated withMYH9disorders

European Journal Of Haematology ◽

10.1111/j.1600-0609.2008.01046.x ◽

2008 ◽

Vol 80 (6) ◽

pp. 540-544 ◽

Cited By ~ 14

Author(s):

Shinji Kunishima ◽

Tadashi Matsushita ◽

Motohiro Hamaguchi ◽

Hidehiko Saito

Keyword(s):

Large Deletion ◽

Identification And Characterization

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Identification and characterization of insertion sequence ISRm1 in Rhizobium meliloti JJ1c10

Canadian Journal of Microbiology ◽

10.1139/m87-053 ◽

1987 ◽

Vol 33 (4) ◽

pp. 314-321 ◽

Cited By ~ 10

Author(s):

Roger Wheatcroft ◽

Robert J. Watson

Keyword(s):

High Frequency ◽

Insertion Sequence ◽

Symbiotic Nitrogen Fixation ◽

Rhizobium Meliloti ◽

Base Pairs ◽

Target Region ◽

Meliloti Strain ◽

Nodc Gene ◽

Identification And Characterization

ISRm1, an insertion sequence present in Rhizobium meliloti strain 1021, has been identified as the cause of the Nod− phenotypes in two mutants of another strain, JJ1c10. The insertions were found to be at different sites, though only about 100 base pairs apart within the nodC gene. ISRm1 causes no mutations in the nifHDK gene region of strain JJ1c10, as it does at high frequency in strain 1021. In JJ1c10 ISRm1 inserted at high frequency into a region of the genome adjacent to copies of other reiterated DNA segments. The target region was not required for symbiotic nitrogen fixation.

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Characterization of the argA Gene Required for Arginine Biosynthesis and Syringomycin Production by Pseudomonas syringae pv. syringae

Applied and Environmental Microbiology ◽

10.1128/aem.69.12.7273-7280.2003 ◽

2003 ◽

Vol 69 (12) ◽

pp. 7273-7280 ◽

Cited By ~ 7

Author(s):

Shi-En Lu ◽

Jonathan D. Soule ◽

Dennis C. Gross

Keyword(s):

Pseudomonas Syringae ◽

Culture Media ◽

Toxin Production ◽

Arginine Biosynthesis ◽

Reading Frame ◽

High Conservation ◽

Plant Pathogenicity ◽

Arginine Deficiency ◽

Dna Fragment

ABSTRACT Two types of necrosis-inducing lipodepsipeptide toxins, called syringomycin and syringopeptin, are major virulence factors of Pseudomonas syringae pv. syringae strain B301D. A previous study showed that a locus, called syrA, was required for both syringomycin production and plant pathogenicity, and the syrA locus was speculated to encode a regulator of toxin production. In this study, sequence analysis of the 8-kb genomic DNA fragment that complements the syrA phenotype revealed high conservation among a broad spectrum of fluorescent pseudomonads. The putative protein encoded by open reading frame 4 (ORF4) (1,299 bp) in the syrA locus region exhibited 85% identity to ArgA, which is involved in arginine biosynthesis in Pseudomonas aeruginosa. Growth of strain W4S2545, the syrA mutant, required supplementation of N minimal medium with arginine. Similarly, syringomycin production of syrA mutant W4S2545 was restored by the addition of arginine to culture media. Furthermore, the insertion of Tn5 in the genome of the syrA mutant W4S2545 was localized between nucleotides 146 and 147 in ORF4, and syringomycin production was complemented in trans with the wild-type DNA fragment containing intact ORF4. These results demonstrate that the syrA locus is the argA gene of P. syringae pv. syringae and that argA is directly involved in arginine biosynthesis and therefore indirectly affects syringomycin production because of arginine deficiency.

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Cloning and characterization of an alternative splicing transcript of the gene coding for human cytidine deaminase

Biochemistry and Cell Biology ◽

10.1139/o06-197 ◽

2007 ◽

Vol 85 (1) ◽

pp. 96-102 ◽

Cited By ~ 1

Author(s):

Bianca Cristina Garcia Lisboa ◽

Tamara da Rocha Machado ◽

Daniel Carvalho Pimenta ◽

Sang Won Han

Keyword(s):

Alternative Splicing ◽

Dna Sequences ◽

Genomic Sequence ◽

Cytidine Deaminase ◽

Alternative Form ◽

Nih3t3 Cells ◽

Reading Frame ◽

Gene Coding ◽

Identification And Characterization

Human cytidine deaminase (HCD) catalyzes the deamination of cytidine or deoxycytidine to uridine or deoxyuridine, respectively. The genomic sequence of HCD is formed by 31 kb with 4 exons and several alternative splicing signals, but an alternative form of HCD has yet to be reported. Here we describe the cloning and characterization of a small form of HCD, HSCD, and it is likely to be a product of alternative splicing of HCD. The alignment of DNA sequences shows that the HSCD matches HCD in 2 parts, except for a deletion of 170 bp. Based on the HCD genome organization, exons 1 and 4 should be joined and all sequences of introns and exons 2 and 3 should be deleted by splicing. This alternative splicing shifted the translation of the reading frame from the point of splicing. The estimated molecular mass is 9.8 kDa, and this value was confirmed by Western blot and mass spectroscopy after expressing the gene fused with glutathionine-S-transferase in the pGEX vector. The deletion and shift of the reading frame caused a loss of HCD activity, which was confirmed by enzyme assay and also with NIH3T3 cells modified to express HSCD and challenged against cytosine arabinoside. In this work we describe the identification and characterization of HSCD, which is the product of alternative splicing of the HCD gene.

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Identification and characterization of G90, a novel mouse RNA that lacks an extensive open reading frame

Gene ◽

10.1016/s0378-1119(99)00120-1 ◽

1999 ◽

Vol 232 (1) ◽

pp. 35-42 ◽

Cited By ~ 9

Author(s):

Ralf Krause ◽

Myriam Hemberger ◽

Heinz Himmelbauer ◽

Vera Kalscheuer ◽

Reinald H. Fundele

Keyword(s):

Open Reading Frame ◽

Reading Frame ◽

Identification And Characterization

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Molecular Cloning and Characterization of cgs, theBrucella abortus Cyclic β(1-2) Glucan Synthetase Gene: Genetic Complementation of Rhizobium meliloti ndvB andAgrobacterium tumefaciens chvB Mutants

Journal of Bacteriology ◽

10.1128/jb.180.17.4392-4400.1998 ◽

1998 ◽

Vol 180 (17) ◽

pp. 4392-4400 ◽

Cited By ~ 54

Author(s):

Nora Iñón de Iannino ◽

Gabriel Briones ◽

Marcelo Tolmasky ◽

Rodolfo A. Ugalde

Keyword(s):

Amino Acids ◽

Membrane Protein ◽

Rhizobium Meliloti ◽

Nodule Development ◽

Nucleotide Sequencing ◽

Insertion Mutant ◽

Reading Frame ◽

Glucan Synthesis ◽

Glucan Synthetase

ABSTRACT The animal pathogen Brucella abortus contains a gene,cgs, that complemented a Rhizobium melilotinodule development (ndvB) mutant and an Agrobacterium tumefaciens chromosomal virulence (chvB) mutant. The complemented strains recovered the synthesis of cyclic β(1-2) glucan, motility, virulence in A. tumefaciens, and nitrogen fixation in R. meliloti; all traits were strictly associated with the presence of an active cyclic β(1-2) glucan synthetase protein in the membranes. Nucleotide sequencing revealed the presence in B. abortus of an 8.49-kb open reading frame coding for a predicted membrane protein of 2,831 amino acids (316.2 kDa) and with 51% identity to R. meliloti NdvB. Four regions of the B. abortus protein spanning amino acids 520 to 800, 1025 to 1124, 1284 to 1526, and 2400 to 2660 displayed similarities of higher than 80% with R. meliloti NdvB. Tn3-HoHo1 mutagenesis showed that the C-terminal 825 amino acids of the Brucella protein, although highly conserved inRhizobium, are not necessary for cyclic β(1-2) glucan synthesis. Confirmation of the identity of this protein as B. abortus cyclic β(1-2) glucan synthetase was done by the construction of a B. abortus Tn3-HoHo1 insertion mutant that does not form cyclic β(1-2) glucan and lacks the 316.2-kDa membrane protein. The recovery of this mutant from the spleens of inoculated mice was decreased by 3 orders of magnitude compared with that of the parental strain; this result suggests that cyclic β(1-2) glucan may be a virulence factor inBrucella infection.

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