Phosphoenolpyruvate Carboxykinase Is an Acid-Induced, Chromosomally Encoded Virulence Factor in Agrobacterium tumefaciens

Abstract

The pckA gene, encoding phosphoenolpyruvate carboxykinase, catalyzes the reversible decarboxylation and phosphorylation of oxaloacetate to form phosphoenolpyruvate. Located on the circular chromosome of Agrobacterium, this locus is adjacent to the loci chvG and chvI, encoding a two-component regulatory system that has been shown to be important in virulence. Using a reporter gene fusion, studies showed that the pckA gene is induced by acidic pH but not by acetosyringone. This acid induction is regulated by the chvG-chvI regulatory system, which controls acid-inducible genes. A pckA mutant had no demonstrable PckA enzyme activity and grew on AB minimal medium with glucose but did not grow on the same medium with succinate as the sole carbon source and was more inhibited in its growth than the wild-type strain by an acidic environment. A pckA mutant was highly attenuated in tumor-inducing ability on tobacco leaf disks and was severely attenuated in vir gene expression. Although vir gene induction was completely restored when a constitutive virG gene was introduced into the mutant strain, virulence was only partially restored. These results suggest that avirulence may be due to a combination of the inhibition of this mutant in the acidic plant wound environment and the poor induction of the vir genes.

Transcriptome Profiling and Functional Analysis ofAgrobacterium tumefaciensReveals a General Conserved Response to Acidic Conditions (pH 5.5) and a Complex Acid-Mediated Signaling Involved inAgrobacterium-Plant Interactions

“…Thus, it is perhaps not surprising that many acid-induced genes are involved in the transport and metabolism of citrate and other members of the TCA cycle, including genes encoding isocitrate lyase (Atu0607), malate:quinone oxidoreductase (Atu0811), sucrose-6-phosphate hydrolase (Atu0944), the TRAP-T family transporter (Atu2470), the TctA subunit of the tricarboxylate transport family (Atu2471), the dicarboxylate MFS transporter (Atu4017), and acetyl-coenzyme A synthase (Atu4130). Consistent with the recent observation that pckA was acid inducible (40), microarray analysis revealed that transcription of pckA was induced around twofold (Table 2). In addition to the organic acids used as a source of carbon and energy, the consumption of organic acids by the TCA cycle results in alkalinization and thus may help maintain internal pH homeostasis (29).…”

“…Microarray analysis revealed that not only does Agrobacterium share general and conserved responses to environmental acid conditions with other bacteria, as described above, but it also has a highly specialized response to perceive the environmental acid as an important signal dedicated to Agrobacterium-plant interactions. In addition to confirming the previously identified acid-inducible genes involved in Agrobacterium-plant interactions, such as aopB, virG, and pckA (13,32,40), the present microarray experiments uncovered many additional acid-inducible genes associated with Agrobacterium-plant interactions. These genes include virE0, virE1, virH1, virH2, and the chvG-chvI system (Table 2).…”

“…This large complement of regulatory elements presumably gives Agrobacterium the ability to sense, respond to, and adapt to a dynamic and changing acidic rhizosphere. In addition to their involvement in induction of the vir regulon, acidic conditions also induce other Agrobacterium determinants required for virulence, such as aopB encoding an outer membrane protein (32) and pckA encoding phosphoenolpyruvate carboxykinase (40). Moreover, salicylic acid, a plant signal important in regulating plant defense, activates the quormone degradation system in Agrobacterium, which also is involved in virulence.…”

Brucella abortus Depends on Pyruvate Phosphate Dikinase and Malic Enzyme but Not on Fbp and GlpX Fructose-1,6-Bisphosphatases for Full Virulence in Laboratory Models

“…abortus 2308 BAB1_2091 is annotated as a pseudogene in some databases (http://www.genome.jp/kegg/) but not in others (http://biocyc.org.). This ORF encodes a protein of 491 amino acids that has 77% similarity to the PckA of the phylogenetically related Agrobacterium tumefaciens, a protein of 536 amino acids known to be functional (35). However, B. abortus BAB1_2091 is separated by a stop codon from an intergenic region that together with BAB1_2090 encodes the last 45 amino acids present in A. tumefaciens PckA.…”

The BatR/BatS Two-Component Regulatory System Controls the Adaptive Response ofBartonella henselaeduring Human Endothelial Cell Infection

“…For the facultative intracellular pathogen Brucella abortus, the BvrS/BvrR TCS is essential for virulence and is responsible for extensive cell envelope modulation, including the upregulation of the outer membrane proteins Omp3a/Omp3b (25,32). The ChvG/ChvI TCS of the plant pathogen Agrobacterium tumefaciens is essential for plant tumor induction (8,33) and controls the expression of acid-inducible genes involved in virulence (29,31). Similarly, the ExoS/ChvI TCS of the legume-nodulating sym-biont Sinorhizobium meliloti plays an essential role in the establishment of symbioses with its host by regulating the production of succinoglycan (11) and was also shown to regulate the expression of genes required for flagellar assembly (57,60).…”