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virG TiBo542

General explanation of virulence

The mechanism for virulence and plant transformation is mostly conserved between A. tumefaciens and A. rhizogenes, with high similarity in the sequences of the virulence genes and their regulation (Moriguchi et al., 2001; Zhu et al., 2000). Therefore, most of the knowledge already available for A. tumefaciens can be extrapolated when working with rhizogenes strains. In fact, the swapping of Ti-plasmids in tumefaciens strains with Ri-plasmids has created some of the most commonly used Agrobacterium rhizogenes strains, including one of the strains used by our team, Arqua1.

The Virulence Mechanism

Agrobacterium strains can transfer large DNA sequences to plant cells and integrate them into the plants' genome. Naturally, all the components for infection are present in a single, non-essential, 250 kbp plasmid (Ti-plasmid in A. tumefaciens or Ri-plasmid in A. rhizogenes).

The genes that code for the mechanism of plant infection and transformation are clustered in the vir (virulence) region, a ~30 kbp region of the Ri-plasmid. There are ca. 35 CDSs distributed in 11 operons in the vir region, which code for - among others - the type IV secretion system (vir B operon), proteins that excise and integrate the T-DNA in the hosts genome (C,D and E operons), and the two-component system that regulates the activation of the whole system (A and G operons). This two-component system can be understood as a “master switch” for the virulence genes.

Vir A is a trans-membrane sensor kinase that reacts to an acidic pH and phenolic compounds secreted by wounded plant tissue, causing it to phosphorylate the response regulator vir G. Among those phenolic compounds are acetosyringone, catechol and vanillin (Bolton et al., 1986). Once phosphorylated, vir G binds to the vir box region (TGAAAT) present in the promoters of virulence operons and upregulates their expression (Aoyama et al., 1989).

pTiBo542

There is a multitude of Agrobacterium strains, with differing characteristics and virulence strengths. The strain A281 in particular, which are able to transform a broader range of plant species and at a higher efficiency, is considered “supervirulent”. Introducing copies of its vir G and vir B operons in regular strains has been shown to recreate the “supervirulent” phenotype. In fact, the virG variant within the pTiBo542 plasmid has been observed to induce a more robust expression of vir genes compared to its virG counterpart on the pTiA6 plasmid. This heightened activity is primarily attributed to the existence of V7I and I106T mutations in the coding sequence of the variant. (Chen et al., 1991).

Sequence and Features