Agrobacterium tumefaciens

***Agrobacterium radiobacter*** (more commonly known as ***Agrobacterium tumefaciens***) is the causal agent of **crown gall disease** (the formation of tumours) in over 140 species of eudicots. It is a rod-shaped, Gram-negative soil bacterium. Symptoms are caused by the insertion of a small segment of DNA (known as the T-DNA, for 'transfer DNA', not to be confused with tRNA that transfers amino acids during protein synthesis), from a plasmid into the plant cell, which is incorporated at a semi-random location into the plant genome. Plant genomes can be engineered by use of *Agrobacterium* for the delivery of sequences hosted in T-DNA binary vectors.

## Conjugation

To be virulent, the bacterium contains tumour-inducing plasmid (Ti plasmid or pTi), of 200 kbp, which contains the T-DNA and all the genes necessary to transfer it to the plant cell. Many strains of *A. tumefaciens* do not contain a pTi.

Since the Ti plasmid is essential to cause disease, prepenetration events in the rhizosphere occur to promote bacterial conjugation - exchange of plasmids amongst bacteria. In the presence of opines, *A. tumefaciens* produces a diffusible conjugation signal called 30C8HSL or the *Agrobacterium* autoinducer[*citation needed*]. This activates the transcription factor TraR, positively regulating the transcription of genes required for conjugation

## Infection methods

*Agrobacterium tumefaciens* infects the plant through its Ti plasmid. The Ti plasmid integrates a segment of its DNA, known as T-DNA, into the chromosomal DNA of its host plant cells. *A. tumefaciens* has flagella that allow it to swim through the soil towards photoassimilates that accumulate in the rhizosphere around roots. Some strains may chemotactically move towards chemical exudates from plants, such as acetosyringone and sugars, which indicate the presence of a wound in the plant through which the bacteria may enter. Phenolic compounds are recognised by the VirA protein, a transmembrane protein encoded in the virA gene on the Ti plasmid. Sugars are recognised by the chvE protein, a chromosomal gene-encoded protein located in the periplasmic space.

At least 25 *vir* genes on the Ti plasmid are necessary for tumor induction. In addition to their perception role, *virA* and *chvE* induce other *vir* genes. The VirA protein has autokinase activity: it phosphorylates itself on a histidine residue. Then the VirA protein phosphorylates the VirG protein on its aspartate residue. The virG protein is a cytoplasmic protein produced from the *virG* Ti plasmid gene. It is a transcription factor, inducing the transcription of the *vir* operons. The ChvE protein regulates the second mechanism of the *vir* genes' activation. It increases VirA protein sensitivity to phenolic compounds.

Attachment is a two-step process. Following an initial weak and reversible attachment, the bacteria synthesize cellulose fibrils that anchor them to the wounded plant cell to which they were attracted. Four main genes are involved in this process: *chvA*, *chvB*, *pscA*, and *att*. The products of the first three genes apparently are involved in the actual synthesis of the cellulose fibrils. These fibrils also anchor the bacteria to each other, helping to form a microcolony.

VirC, the most important virulent protein, is a necessary step in the recombination of illegitimate recolonization. It selects the section of the DNA in the host plant that will be replaced and it cuts into this strand of DNA.

## Genes in the T-DNA

### Hormones

To cause gall formation, the T-DNA encodes genes for the production of auxin or indole-3-acetic acid via the IAM pathway. This biosynthetic pathway is not used in many plants for the production of auxin, so it means the plant has no molecular means of regulating it and auxin will be produced constitutively. Genes for the production of cytokinins are also expressed. This stimulates cell proliferation and gall formation.