Unleashing Nature’s Genetic Engineer: The Remarkable Role of Agrobacterium Tumefaciens in Plant Science

Plant genetic transformation is a significant technical development in modern science that has not only enabled basic insights into plant biology but has also heralded a new age in crop improvement and commercial farming. However, advancements in plant tissue culture technologies are required to streamline operational stages to enhance transformation efficiency. Furthermore, substantial emphasis must be placed on uncovering genes that influence developmental reprogramming and homologous recombination, as this information will aid in better understanding their function in boosting the efficiency of genetic transformation in plants¹.

Agrobacterium tumefaciens, used for genetic transformation, was first isolated from grapevine galls in 1897². A. tumefaciens is a ubiquitous soil bacterium that can live independently or as a pathogen in association with a plant host. Its virulence proficiency is dependent on the presence of the Tumour-inducing (Ti) plasmid. A. tumefaciens has two direct repeat sequences of about 25 base pairs that make up its transferred DNA (T-DNA). Most of the research has used either the octopine-using strain containing pTiA6 or the nopaline-metabolizing strains containing plasmids pTiC58 and pTiT37, respectively. When living independently, the A. tumefaciens virulence effect is not activated. A. tumefaciens activates its chromosomal virulence genes (chv genes) and plasmid encoded virulence genes (vir genes) in response to plant-derived cues in the rhizosphere. Vir genes are directly engaged in T-DNA cleavage from the Ti plasmid, T-DNA processing, transferring, and integration into plant nuclei, whereas chv genes play significant roles in A. tumefaciens pathogenicity signal transduction. T-DNA encodes genes that produce indole-3-acetic acid (IAA) and cytokinin (CK). These lead to the production of many plant hormones that promote uncontrolled cell division and undifferentiated growth of plant tissues. This situation results in the formation of a plant tumour and permanent plant genetic transformation. Oncogenic genes expressed by T-DNA are neither physically nor physiologically necessary for T-DNA transmission. T-DNA encoded genes can thus be eliminated and replaced with desired genes. “T-DNA” that has been genetically altered can still be transported, incorporated, and expressed in plant cells³.

Plant transfer of transformed Agrobacterium tumefaciens was easily achieved by the flower dipping method. This method was tested on Arabidopsis developing flowering tissues. Transformed progeny were obtained by immersing these tissues in a solution of Agrobacterium tumefaciens, 500 microliters per liter of surfactant Silwet L-77 and 5% sucrose⁴.

A multicellular population of one or more microorganisms that is firmly attached to a surface and frequently covered in an extracellular matrix of secreted biopolymers is referred to as a biofilm⁵. By transferring and integrating a section of transferred DNA (T-DNA) encoded by a plasmid into the host genome, the bacterium A. tumefaciens genetically alters plant cells. In vitro, A. tumefaciens adheres to and develops a sophisticated biofilm on a range of biotic and abiotic substrates.  A sophisticated regulatory network that controls surface attachment, motility, and cell division gives the A. tumefaciens life cycle an unexpected asymmetry. A. tumefaciens pathogenicity is associated with biofilm development both mechanistically and via common regulatory molecules. The second messenger cyclic-di-GMP, nutrient concentrations, and the function of the plant host are just a few of the internal and external (environmental) elements that influence the development of biofilm state⁶.

References:

1. Ramkumar TR, Lenka SK, Arya SS, Bansal KC. A short history and perspectives on plant genetic transformation. In: Methods and Protocols in Biolistic DNA Delivery in Plants. 2020;39-68.
2. Cavara F. Eziologia di alcune malattie di piante cultivate. Le Stazioni Sperimentale Agraric Itliana. 1897;30:482-509.
3. Subramoni S, Nathoo N, Klimov E, Yuan ZC. Agrobacterium tumefaciens responses to plant-derived signaling molecules. Frontiers in plant science. 2014;5:322
4. Clough, S. J., & Bent, A. F. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. The Plant Journal. 1998;16(6):735-743.
5. Costerton, J. W., Lewandowski, Z., Caldwell, D. E., Korber, D. R., and Lappin-Scott, H. M. (1995). Microbial biofilms.  Rev. Microbiol.49, 711–745.Doi: 10.1146/annurev.mi.49.100195.00343
6. Heindl JE, Wang Y, Heckel BC, Mohari B, Feirer N, Fuqua C. Mechanisms and regulation of surface interactions and biofilm formation in Agrobacterium. Frontiers in Plant Science. 2014;5:176

Figure References:

1. GAO, Caixia. Genome engineering for crop improvement and future agriculture. Cell, 2021, 184.6: 1621-1635. Receive Date: 16.05.2023, https://www.sciencedirect.com/science/article/pii/S0092867421000052
2. Adriana Galloge. Into Agrobacterium and Plasmıds . Receive Date: 16.05.2023, https://goldbio.com/articles/article/Intro-Agrobacterium-and-Plasmids
3. Heindl JE, Wang Y, Heckel BC, Mohari B, Feirer N, Fuqua C. Mechanisms and regulation of surface interactions and biofilm formation in Agrobacterium. Frontiers in Plant Science. 2014;5:176. Receive Date: 16.05.2023, https://www.frontiersin.org/articles/10.3389/fpls.2014.00176/full

Inspector:Elif BÖCÜ