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Plant pathogenic bacteria, such as Pseudomonas syringae pv. tomato strain DC3000, the causative agent of tomato bacterial speck disease, grow to high levels in the apoplastic space between plant cells. Colonization of plant tissue requires expression of virulence factors that modify the apoplast to make it more suitable for pathogen growth or facilitate adaptation of the bacteria to the apoplastic environment. To identify new virulence factors involved in these processes, DC3000 Tn5 transposon insertion mutants with reduced virulence on Arabidopsis thaliana were identified. In one of these mutants, the Tn5 insertion disrupted the malate:quinone oxidoreductase gene (mqo), which encodes an enzyme of the tricarboxylic acid cycle, mqo mutants do not grow to wild-type levels in plant tissue at early time points during infection. Further, plants infected with mqo mutants develop significantly reduced disease symptoms, even when the growth of the mqo mutant reaches wild-type levels at late stages of infection. Mutants lacking mqo function grow more slowly in culture than wild-type bacteria when dicarboxylates are the only available carbon source. To explore whether dicarboxylates are important for growth of DC3000 in the apoplast, we disrupted the dctA1 dicarboxylate transporter gene. DC3000 mutants lacking dctA1 do not grow to wild-type levels in planta, indicating that transport and utilization of dicarboxylates are important for virulence of DC3000. Thus, mqo may be required by DC3000 to meet nutritional requirements in the apoplast and may provide insight into the mechanisms underlying the important, but poorly understood process of adaptation to the host environment.

The high oxygen affinity of Ascaris hemoglobin domain I (D1) appears to be associated with its structural similarity to myoglobin. The x-ray-derived crystal structure shows a strong hydrogen bond between tyrosine at position 10 in the B helix of D1 and the distal oxygen of the ligand, in combination with a weak hydrogen bond between glutamine-E7 and the proximal oxygen. These bonds hold the ligand in the binding pocket. A third hydrogen bond between these two amino acids gives stability to the structure.

AvrRpt2, a Pseudomonas syringae type III effector protein, functions from inside plant cells to promote the virulence of P. syringae pv. tomato strain DC3000 ( PstDC3000) on Arabidopsis thaliana plants lacking a functional copy of the corresponding RPS2 resistance gene. In this study, we extended our understanding of AvrRpt2 virulence activity by exploring the hypothesis that AvrRpt2 promotes PstDC3000 virulence by suppressing plant defenses. When delivered by PstDC3000, AvrRpt2 suppresses pathogen-related ( PR) gene expression during infection, suggesting that AvrRpt2 suppresses defenses mediated by salicylic acid (SA). However, AvrRpt2 promotes PstDC3000 growth on transgenic plants expressing the SA-degrading enzyme NahG, indicating that AvrRpt2 does not promote bacterial virulence by modulating SA levels during infection. AvrRpt2 general virulence activity does not depend on the RPM1 resistance gene, as mutations in RPM1 had no effect on AvrRpt2-induced phenotypes. Transgenic plants expressing AvrRpt2 displayed enhanced susceptibility to PstDC3000 strains defective in type III secretion, indicating that enhanced susceptibility of these plants is not because of suppression of defense responses elicited by other type III effectors. Additionally, avrRpt2 transgenic plants did not exhibit increased susceptibility to Peronospora parasitica and Erysiphe cichoracearum, suggesting that AvrRpt2 virulence activity is specific to P. syringae. [ABSTRACT FROM AUTHOR]

To identify virulence genes of P. syringae pv. tomato strain DC3000 we screened for mutants with reduced virulence on its plant hosts, Arabidopsis thaliana and tomato. We isolated a Tn5-insertion mutant that exhibited reduced virulence on both hosts. Further characterization showed that this mutant carried a single Tn5 insertion in the dsbA gene, which encodes a periplasmic disulphide bond-forming protein. In addition to reduced virulence, the dsbA mutant exhibits mucoid colony morphology, Ioss of fluorescence, decreased motility, and a reduced growth rate in culture. The dsbA mutant is able to multiply in A. thaliana and tomato plants, trigger the hypersensitive response on tobacco and elicit Pto-mediated resistance in tomato, indicating that type III secretion occurs in this background. However, type III secretion appears to function with reduced efficiency in the dsbA mutant, as type III-dependent secretion of HrpZ and AvrRpt2 is impaired. These findings indicate that while the dsbA gene is required for multiple cellular functions in P. syringae, type III secretion in P. syringae is only partially dependent on dsbA. [ABSTRACT FROM AUTHOR]