The world of GMOs: from discovery to analysis (part 2)

History of GMOs: agrobacterium tumefaciens

The third and final piece in the history of GMOs concerns A. tumefaciens. This gram-positive bacterium is capable of mainly infecting dicotyledons and causing growth in them comparable to that of a tumor.

Crown gall tumor, plant tumor, World of OGMs
Figure 1 – Crown gall tumor. [Source: jacilluch– flickr.com]

Specifically, when a plant is injured and its resistant surface is damaged, through these less protected areas A. tumefaciens will be able to penetrate the plant tissue and transfer a piece of its DNA to the chromosome of the plant cell.

In this way, the plant cell will become a tumor, starting an uncontrolled growth that will generate the pathological condition.

Galla del colletto; tumor plant
A. tumefaciens
Figure 2 – Tumor growth.  [Source: microbiologiaitalia]

Genesis of tumor growth

Various factors are involved in the mechanism of the formation of this particular plant tumor. First of all, following a wound, a plant releases unpleasant and repellent phenolic compounds which act as attractors for A. tumefaciens. In this way, this bacterium recognizes the injured and most attackable areas of the plant.

Specifically, these compounds bind surface bacterial receptors that trigger signal transduction with phosphorylation of a protein, which then induces the transcription of Vir genes that will produce various Vir proteins. In particular, we are talking about SSB (single-strand binding protein) proteins. In particular, we distinguish:

  • VirD protein responsible for DNA double cutting at two sites of the Ti (tumor inducing) plasmid that will induce the tumor. In fact, starting from the latter, a fragment of T-DNA (transforming DNA) will detach as a single helix;
  • Vir E which intervenes to protect the single strand of T-DNA from degradation;
  • Vir B which will form a real channel between the bacterial cell and the plant one, acting as a bridge and connecting the two membranes between them.

Then, through this “bridge” the T-DNA fragment, respectively bound to the SSB proteins, will be transferred into the plant cell. At this point, once the transfer has taken place, the T-DNA from the cytoplasm of the plant cell enters the nucleus and is integrated into the plant genome.

Genesi della galla del colletto, genesis of tumor plant; GMOs world A. tumefaciens
Figure 3 – Genesis of tumor growth [Source: docplayer]

The integration into chromosomes is specific and occurs in the vicinity of the opines, genes responsible for regulating particular growth factors of plant cells; these genes cause the cell to start duplicating itself in an uncontrolled way, developing the tumor.

Gene editing and A. tumefaciens

A. tumefaciens, therefore, constitutes a tool for introducing specific bacterial DNA into the genome of a plant cell, which in this case is T-DNA.

Meccanismo d'inserzione genica dell'A. tumefaciens, genetics insertion of A. tumefaciens
Figure 4 – gene insertion mechanism of A. tumefaciens  [Source: slideplayer]

The scholars who first created GMOs imagined inserting, instead of T-DNA, a DNA fragment of interest (such as that of the bt toxin gene), thus providing the plant considered with an exogenous gene. In doing so, we came to the production of transgenic plants.

Specifically, by introducing the bt gene and making the plant cell proliferate, an entire plant is generated which in each of its cells produces the bt toxin, the so-called self-protected plant. Therefore, as soon as the borer larva begins to eat the leaves of this type of plant, ingesting the BT toxin, it immediately dies without having time to cause damage to the plantation.

Therefore, by exploiting the knowledge acquired based on these three elements:

  • bt toxin;
  • restriction enzymes;
  • gene insertion mechanism of A. tumefaciens.

We came to the generation of the first generation GMO plants. With this procedure, many modified plants capable of growing in extremely arid soils are also created today.

Second generation GMOs: from nuclear to plastid transformation

This type of transformation no longer involves bacteria because A. tumefaciens is able to transfer DNA only at the nuclear level.

In fact, in this case, the transformative process is mechanical and takes place by adhering the transgenic DNA to tungsten or gold micro-bullets.

Subsequently, through a real micro-cannon, these particles conjugated to the transgenic DNA will be “fired” on the target cells.

Thus, this DNA will enter the nucleus x times and the chloroplasts or mitochondria y times. At this point, a screening of the cells transformed at the plastid level is carried out. These cells will then be used to create a new second-generation GMO transgenic plant.

The characteristic of this type of plant is that it does not spread the transgenic character to other plantations; in fact, chloroplasts are semi-autonomous organelles that have their DNA. Therefore, by exclusively carrying out the GMO transformation of the plastid DNA of the male gametes, the diffusion of the transgenic DNA will not take place.

Positive and negative aspects of GMOs

Contrary to the widespread general ideology, the creation of GMOs is to be considered with a positive meaning as there have been very few negative cases.

In particular, among the various cases of incorrect use of GMOs we remember, for example, that due to particular fatty acids of the Brazilian walnut that are very allergenic for some people, which is why their production was stopped.

The strange case of the “strawberry fish”

In an attempt to discredit the use of GMOs, real “hoaxes” have emerged over the years, including the case of the famous “strawberry fish” that has never been marketed or even made.

La famosa "fragola pesce", strawberry fish
Figure 5 – the famous “strawberry fish” [Source: ilfattoquotidiano]

To understand this particular case assembled ad hoc, first of all, we understand why it could be interesting to create this type of GMO.

Among the various transgenics, some have a particular type of tolerance to cold shock or cold stress. This stress is a big problem for the cell because, for example, a cell that perhaps lives very well at 30 ° C could risk dying at 10 ° C.

This is because the cold shock acts by causing the formation of secondary structures that make the translation of mRNA impossible, not allowing the production of some proteins. Since it is necessary to have continuous production of many proteins if this were to fail the body would go to death.

One of the fruits particularly damaged by sudden drops in temperature is precisely the strawberry.

Csp protein

To solve the problem of the formation of secondary structures of mRNA, in nature there are proteins, the csp (cold shock proteins) that go to bind the regions of the messenger.

In particular, strawberry has been found to lack these proteins. Therefore, it was hypothesized to introduce the csp genes into the genome of the aforementioned false fruit. In this regard, it was thought to use those coming from the genome of Antarctic fish, specifically from cod.

This modification has never been made but from the simple idea, from the simple attempt, a false myth arose in the collective imagination that greatly damaged the reputation of GMOs.

Indeed, in Italy this demonization has led to a moratorium against GMOs, in particular those used in agriculture, so today we find ourselves in a paradoxical situation: we import GMO fruits and vegetables from abroad, we use them, we eat them, we buy them. , but by law, we cannot produce it.

GMO and virus resistance

Through the GMO technique, it is also possible to create plants resistant to viruses which have always been a real plague for many agricultural productions.

Specifically, most viruses need an entrance door to infect the host, often consisting of a membrane receptor.

What can be done is to generate a modified receptor so that the domain of the natural ligand continues to be functional, but the contact area of the virus will no longer be compatible with viral attachment.

Through this modification, the organism that is created is not transgenic because it simply modifies a gene already possessed by a particular cell, carrying out a so-called in vitro evolution.

Giovanna Spinosa

translation of the article by: Umberto Lazzaro

Sources

  • https://www.microbiologiaitalia.it/didattica/organismi-geneticamente-modificati-ogm/;
  • https://kids.frontiersin.org/articles/10.3389/frym.2020.00064;
  • https://www.researchgate.net/profile/Girma-Alelign/publication/347240660_The_Effects_of_Genetically_Modified_Organisms_GMO_on_Environment_and_Molecular_Techniques_to_Minimize_Its_Risk/links/5fe6348492851c13febda77e/The-Effects-of-Genetically-Modified-Organisms-GMO-on-Environment-and-Molecular-Techniques-to-Minimize-Its-Risk.pdf.

Image credits

  • Figure 1: https://www.flickr.com/photos/70626035@N00/6896084714/;
  • Figure 2: https://www.microbiologiaitalia.it/batteriologia/agrobacterium-tumefaciens-il-batterio-che-sfrutta-le-piante/;
  • Image 3: https://docplayer.it/54011228-Ingegneria-genetica-favorevoli-o-contrari.html;
  • Image 4: https://slideplayer.it/slide/954101/;
  • Figure 5: https://www.ilfattoquotidiano.it/2012/05/02/dice-report/215103/.

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