scholarly journals PhyA, a Secreted Protein of Xanthomonas oryzae pv. oryzae, Is Required for Optimum Virulence and Growth on Phytic Acid as a Sole Phosphate Source

2003 ◽  
Vol 16 (11) ◽  
pp. 973-982 ◽  
Author(s):  
Subhadeep Chatterjee ◽  
Rajan Sankaranarayanan ◽  
Ramesh V. Sonti
Keyword(s):  
Phytic Acid ◽  
Xanthomonas Campestris ◽  
Xanthomonas Oryzae ◽  
Bacillus Species ◽  
Active Site Residues ◽  
Significant Similarity ◽  
Reading Frame ◽  
Predicted Signal Peptide ◽  
Serious Disease ◽  
Phosphate Source

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. We have identified a novel virulence deficient mutant (BXO1691) of X. oryzae pv. oryzae that has a Tn5 insertion in an open reading frame (phyA; putative phytase A) encoding a 373-amino acid (aa) protein containing a 28-aa predicted signal peptide. Extracellular protein profiles revealed that a 38-kDa band is absent in phyA mutants as compared with phyA+ strains. A BLAST search with phyA and its deduced polypeptide sequence indicated significant similarity with conserved hypothetical proteins in Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris and limited homology to secreted phytases of Bacillus species. Homology modeling with a Bacillus phytase as the template suggests that the PhyA protein has a similar six-bladed β-propeller architecture and exhibits conservation of certain critical active site residues. Phytases are enzymes that are involved in degradation of phytic acid (inositol hexaphosphate), a stored form of phosphate in plants. The phyA mutants exhibit a growth deficiency in media containing phytic acid as a sole phosphate source. Exogenous phosphate supplementation promotes migration of phyA X. oryzae pv. oryzae mutants in rice leaves. These results suggest that the virulence deficiency of phyA mutants is, at least in part, due to inability to use host phytic acid as a source of phosphate. phyA-like genes have not been previously reported to be involved in the virulence of any plant pathogenic bacterium.

rpfF Mutants of Xanthomonas oryzae pv. oryzae Are Deficient for Virulence and Growth Under Low Iron Conditions

2002 ◽  
Vol 15 (5) ◽  
pp. 463-471 ◽  
Author(s):  
Subhadeep Chatterjee ◽  
Ramesh V. Sonti
Keyword(s):  
Xanthomonas Campestris ◽  
Iron Supplementation ◽  
Extracellular Enzymes ◽  
Phenotypic Expression ◽  
Tetracycline Resistance ◽  
Extracellular Enzyme ◽  
Extracellular Polysaccharide ◽  
Xanthomonas Oryzae ◽  
Uptake System ◽  
Serious Disease

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. In the related bacterium Xanthomonas campestris pv. campestris, the rpfF gene is involved in production of a diffusible extracellular factor (DSF) that positively regulates synthesis of virulence-associated functions like extracellular polysaccharide (EPS) and extracellular enzymes. Transposon insertions in the rpfF homolog of X. oryzae pv. oryzae are deficient for virulence and production of a DSF but are proficient for EPS and extracellular enzyme production. The rpfF X. oryzae pv. oryzae mutants exhibit an unusual tetracycline susceptibility phenotype in which exogenous iron supplementation is required for phenotypic expression of a tetracycline resistance determinant that is encoded on an introduced plasmid. The rpfF X. oryzae pv. oryzae mutants also overproduce one or more siderophores and exhibit a growth deficiency under low iron conditions as well as in the presence of reducing agents that are expected to promote the conversion of Fe+3 to Fe+2. Exogenous iron supplementation promotes migration of rpfF X. oryzae pv. oryzae mutants in rice leaves. The results suggest that rpfF may be involved in controlling an iron-uptake system of X. oryzae pv. oryzae and that an inability to cope with the conditions of low iron availability in the host may be the reason for the virulence deficiency of the rpfF X. oryzae pv. oryzae mutants.

scholarly journals XadM, a Novel Adhesin of Xanthomonas oryzae pv. oryzae, Exhibits Similarity to Rhs Family Proteins and Is Required for Optimum Attachment, Biofilm Formation, and Virulence

2012 ◽  
Vol 25 (9) ◽  
pp. 1157-1170 ◽  
Author(s):  
Binod B. Pradhan ◽  
Manish Ranjan ◽  
Subhadeep Chatterjee
Keyword(s):  
Pathogenic Bacteria ◽  
Surface Protein ◽  
Growth Conditions ◽  
Xanthomonas Oryzae ◽  
Carbohydrate Binding ◽  
Significant Similarity ◽  
Reading Frame ◽  
Family Protein ◽  
Bacterial Blight Pathogen

By screening a transposon-induced mutant library of Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, we have identified a novel 5.241-kb open reading frame (ORF) named xadM that is required for optimum virulence and colonization. This ORF encodes a protein, XadM, of 1,746 amino acids that exhibits significant similarity to Rhs family proteins. The XadM protein contains several repeat domains similar to a wall-associated surface protein of Bacillus subtilis, which has been proposed to be involved in carbohydrate binding. The role of XadM in X. oryzae pv. oryzae adhesion was demonstrated by the impaired ability of an xadM mutant strain to attach and form biofilms. Furthermore, we show that XadM is exposed on the cell surface and its expression is regulated by growth conditions and plays an important role in the early attachment and entry inside rice leaves. Interestingly, XadM homologs are present in several diverse bacteria, including many Xanthomonas spp. and animal-pathogenic bacteria belonging to Burkholderia spp. This is the first report of a role for XadM, an Rhs family protein, in adhesion and virulence of any pathogenic bacteria.

Virulence deficiency caused by a transposon insertion in the purH gene of Xanthomonas oryzae pv. oryzae

2005 ◽  
Vol 51 (7) ◽  
pp. 575-581 ◽  
Author(s):  
Subhadeep Chatterjee ◽  
Ramesh V Sonti
Keyword(s):  
Xanthomonas Campestris ◽  
Pathogenic Bacteria ◽  
Minimal Medium ◽  
Pathogen Resistance ◽  
Xanthomonas Oryzae ◽  
Chromosomal Dna ◽  
Transposon Insertion ◽  
Growth Deficiency ◽  
Related Plant ◽  
Serious Disease

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. We have identified a Tn5-induced virulence-deficient mutant (BXO1704) of X. oryzae pv. oryzae. The BXO1704 mutant exhibited growth deficiency in minimal medium but was proficient in inducing a hypersensitive response in a non-host tomato plant. Sequence analysis of the chromosomal DNA flanking the Tn5 insertion indicated that the Tn5 insertion is in the purH gene, which is highly homologous to purH genes of other closely related plant pathogenic bacteria Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris. Purine supplementation reversed the growth deficiency of BXO1704 in minimal medium. These results suggest that the virulence deficiency of BXO1704 may be due to the inability to use sufficient purine in the host.Key words: auxotroph, plant pathogen, resistance.

scholarly journals Mutants of Xanthomonas oryzae pv. oryzae Deficient in General Secretory Pathway Are Virulence Deficient and Unable to Secrete Xylanase

2000 ◽  
Vol 13 (4) ◽  
pp. 394-401 ◽  
Author(s):  
Suvendra K. Ray ◽  
R. Rajeshwari ◽  
Ramesh V. Sonti
Keyword(s):  
Xanthomonas Campestris ◽  
Secretory Pathway ◽  
Genomic Library ◽  
Xanthomonas Oryzae ◽  
Enzyme Assays ◽  
Specific Primers ◽  
Xylanase Production ◽  
Protein Secretion System ◽  
Serious Disease ◽  
Xanthomonas Campestris Pv Campestris

Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial leaf blight, a serious disease of rice. A virulence- and xylanase-deficient mutant of Xoo was isolated following ethyl methane sulfonate (EMS) mutagenesis. A cosmid clone that restored virulence and xylanase secretion was obtained from a genomic library by functional complementation. Trans-poson mutagenesis and marker exchange studies revealed genes on the cloned DNA that were required for xylanase production and virulence. Sequence analysis with trans-poson-specific primers revealed that these genes were homologues of xps F and xps D, which encode components of a protein secretion system in Xanthomonas campestris pv. campestris. Enzyme assays showed xylanase accumulation in the periplasmic space and cytoplasm of the xps F mutant and the complementing clone restored transport to the extracellular space.

scholarly journals Isolation and characterization of PEP3, a gene required for vacuolar biogenesis in Saccharomyces cerevisiae

1991 ◽  
Vol 11 (12) ◽  
pp. 5801-5812
Author(s):  
R A Preston ◽  
M F Manolson ◽  
K Becherer ◽  
E Weidenhammer ◽  
D Kirkpatrick ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Product ◽  
Zinc Finger ◽  
Genomic Library ◽  
Sequence Similarity ◽  
Carboxypeptidase Y ◽  
Lag Phase ◽  
Significant Similarity ◽  
Reading Frame ◽  
Isolation And Characterization

The Saccharomyces cerevisiae PEP3 gene was cloned from a wild-type genomic library by complementation of the carboxypeptidase Y deficiency in a pep3-12 strain. Subclone complementation results localized the PEP3 gene to a 3.8-kb DNA fragment. The DNA sequence of the fragment was determined; a 2,754-bp open reading frame predicts that the PEP3 gene product is a hydrophilic, 107-kDa protein that has no significant similarity to any known protein. The PEP3 predicted protein has a zinc finger (CX2CX13CX2C) near its C terminus that has spacing and slight sequence similarity to the adenovirus E1a zinc finger. A radiolabeled PEP3 DNA probe hybridized to an RNA transcript of 3.1 kb in extracts of log-phase and diauxic lag-phase cells. Cells bearing pep3 deletion/disruption alleles were viable, had decreased levels of protease A, protease B, and carboxypeptidase Y antigens, had decreased repressible alkaline phosphatase activity, and contained very few normal vacuolelike organelles by fluorescence microscopy and electron microscopy but had an abundance of extremely small vesicles that stained with carboxyfluorescein diacetate, were severely inhibited for growth at 37 degrees C, and were incapable of sporulating (as homozygotes). Fractionation of cells expressing a bifunctional PEP3::SUC2 fusion protein indicated that the PEP3 gene product is present at low abundance in both log-phase and stationary cells and is a vacuolar peripheral membrane protein. Sequence identity established that PEP3 and VPS18 (J. S. Robinson, T. R. Graham, and S. D. Emr, Mol. Cell. Biol. 11:5813-5824, 1991) are the same gene.

scholarly journals Mutations in the Predicted Active Site of Xanthomonas oryzae pv. oryzae XopQ Differentially Affect Virulence, Suppression of Host Innate Immunity, and Induction of the HR in a Nonhost Plant

2015 ◽  
Vol 28 (2) ◽  
pp. 195-206 ◽  
Author(s):  
Mahesh Kumar Gupta ◽  
Rajkanwar Nathawat ◽  
Dipanwita Sinha ◽  
Asfarul S. Haque ◽  
Rajan Sankaranarayanan ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Active Site ◽  
Plant Cell Wall ◽  
Structural Model ◽  
Bioinformatic Analysis ◽  
Xanthomonas Oryzae ◽  
Active Site Residues ◽  
Callose Deposition ◽  
Nonhost Plant ◽  
Bacterial Blight Pathogen

Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, secretes a number of effectors through a type 3 secretion system. One of these effectors, called XopQ, is required for virulence and suppression of rice innate immune responses induced by the plant cell-wall-degrading enzyme lipase/esterase A (LipA). Bioinformatic analysis suggested that XopQ is homologous to inosine-uridine nucleoside hydrolases (NH). A structural model of XopQ with the protozoan Crithidia fasciculata purine NH suggested that D116 and Y279 are potential active site residues. X. oryzae pv. oryzae xopQ mutants (xopQ−/pHM1::xopQD116A and xopQ−/pHM1::xopQY279A) show reduced virulence on rice compared with xopQ−/pHM1::xopQ. The two predicted XopQ active site mutants (xopQ−/pHM1::xopQD116A and xopQ−/pHM1::xopQY279A) exhibit a reduced hypersensitive response (HR) on Nicotiana benthamiana, a nonhost. However, Arabidopsis lines expressing either xopQ or xopQY279A are equally proficient at suppression of LipA-induced callose deposition. Purified XopQ does not show NH activity on standard nucleoside substrates but exhibits ribose hydrolase activity on the nucleoside substrate analogue 4-nitrophenyl β-D-ribofuranoside. The D116A and Y279A mutations cause a reduction in biochemical activity. These results indicate that mutations in the predicted active site of XopQ affect virulence and induction of the HR but do not affect suppression of innate immunity.

scholarly journals Novel Genomic Locus with Atypical G+C Content that Is Required for Extracellular Polysaccharide Production and Virulence in Xanthomonas oryzae pv. oryzae

2001 ◽  
Vol 14 (11) ◽  
pp. 1335-1339 ◽  
Author(s):  
Sridhar Dharmapuri ◽  
J. Yashitola ◽  
M. R. Vishnupriya ◽  
Ramesh V. Sonti
Keyword(s):  
Sequence Data ◽  
Extracellular Polysaccharide ◽  
Genomic Region ◽  
Xanthomonas Oryzae ◽  
Genomic Locus ◽  
Reading Frame ◽  
Glycosyl Transferase ◽  
Clone Sequence ◽  
Gum Gene Cluster ◽  
Transposon Insertions

Three exopolysaccharide (EPS)- and virulence-deficient mutants of Xanthomonas oryzae pv. oryzae, the causal agent of bacterial leaf blight of rice, were isolated by Tn5 mutagenesis. These insertions are not located within the gum gene cluster. A 40-kb cosmid clone that restored EPS production and virulence to all three mutants was isolated, and the three transposon insertions were localized to contiguous 4.3- and 3.5-kb EcoRI fragments that are included in this clone. Sequence data indicate that two of the transposon insertions are in genes that encode a putative sugar nucleotide epimerase and a putative glycosyl transferase, respectively; the third insertion is located between the glycosyl transferase gene and a novel open reading frame (ORF). A 5.5-kb genomic region in which these three ORFs are located has a G+C content of 51.7%, quite different from the G+C content of approximately 65.0% that is typical of X. oryzae pv. oryzae. Homologues of this locus have not yet been reported in any other xanthomonad.

Characterization of the African swine fever virus protein p49: a new late structural polypeptide

Microbiology ◽  
2000 ◽  
Vol 81 (1) ◽  
pp. 59-65 ◽  
Author(s):  
Inmaculada Galindo ◽  
Eladio Viñuela ◽  
Angel L. Carrascosa
Keyword(s):  
Cell Attachment ◽  
African Swine Fever Virus ◽  
Rabbit Antiserum ◽  
African Swine Fever ◽  
Virus Genome ◽  
Fever Virus ◽  
Virus Protein ◽  
Significant Similarity ◽  
Reading Frame ◽  
Cell Extracts

The open reading frame B438L, located within the EcoRI B fragment of the African swine fever virus genome, is predicted to encode a protein of 438 amino acids with a molecular mass of 49·3 kDa. It presents a cell attachment RGD (Arg–Gly–Asp) motif but no other significant similarity to protein sequences in databases. Northern blot and primer extension analysis showed that B438L is transcribed only at late times during virus infection. The B438L gene product has been expressed in Escherichia coli, purified and used as an antigen for antibody production. The rabbit antiserum specific for pB438L recognized a protein of about 49 kDa in virus-infected cell extracts. This protein was synthesized late in infection by all the virus strains tested, was located in cytoplasmic virus factories and appeared as a structural component of purified virus particles.

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