The genetic modification methods involving CRISPR have been a part of science and popular culture for over a decade and we have begun to see scientific accomplishments using Cas9 and all the other CRISPR tools. But it has also been a question of when we’d start seeing items such as food modified by CRISPR become a commonplace sight. While we’ve seen plenty of published studies discussing such advancements, including in our very own articles here on Bioscription, that research has yet to progress to public availability. And the foods that have come out, such as arctic apples and innate potatoes, instead were made using RNA interference (RNAi) tools that silenced unwanted genes rather than gene editing changes using CRISPR, as also seen in other research.
This may all be changing with a new tomato that was just released in Japan, referred to as the Sicilian Rouge High GABA tomato. The name is fairly straightforward, being a cultivar of the Sicilian Rouge variety and the tomato being gene edited to have a high amount of the molecule GABA inside of it. Gamma-Aminobutyric acid (GABA) is a neurotransmitter used in animals in general to help regulate stress responses and it has become common to take it orally as a medicine to reduce blood pressure. Tomatoes are already significantly high in GABA as compared to other crops, so are a great starting point to push this capability even further.
While this new product was officially released just the other week as a collaboration between researchers at the University of Tsukuba and the Sanatech Seed company, the origins of this tomato go quite a bit further back from that. The original study from said researchers that proved such a tomato could be made was carried out in 2017 and it is that study that we’re going back to today to see what they did and how their CRISPR changes improved the tomato.
The Hunt For GABA
The promotion of foods high in GABA has been an important goal for medical researchers for years, but has run into issues of food cultures and the usage of only specific foods grown in certain regions. Tomatoes are an exception to this, as they are used in most cuisines around the world and already have a high GABA starting value, making them perfect for increasing consumption habits that result in lower blood pressure. If that GABA production amount can be increased further, it would allow tomatoes to supplant most other methods of having GABA-rich foods in the diet of the general public.
Tomatoes have been found to have a massive storage capacity for GABA, which primarily occurs during the greening stage of fruit development and not later red coloration stages. Earlier testing in 2008 was able to identify three genes involved in making GABA from the amino acid glutamate and were able to be overexpressed using a promoter sequence used during the fruit ripening stage. This increased GABA accumulation in the tomatoes by 600 to 700%. Similarly, other studies using RNAi to silence the specific gene involved in regulating and restricting GABA biosynthesis found an even greater 920% improvement in GABA. Yet further studies found a higher level of biosynthesis after deleting the auto-inhibitory regions of one of the genes so it would always be transcribed and used.
A Chance For CRISPR
While previous attempts by the researchers tried to purposefully make mutations in this auto-inhibitory region to allow GABA accumulation to occur during the entire fruit growth cycle, targeted mutagenesis using chemicals like EMS and using standard tools such as Targeting Induced Local Lesions In Genomes (TILLING) to identify such mutations did not uncover any such mutations despite multiple attempts. Therefore, they turned to CRISPR to directly induce a change to this region in a directed and controlled manner, rather than relying on random mutagenesis to accomplish it.
They focused on using the Cas9 version to create double-strand breaks in the DNA at the desired location and relied upon the non-homologous end joining (NHEJ) repair system to inaccurately stitch the damaged DNA back together with altered nucleotides, thus rendering the auto-inhibitory region nonfunctional. This was done for both of the GABA biosynthesis genes and the scientists were able to observe both deletions of nucleotides at the site and insertions of inaccurate nucleotides into the site. Compared to the control having a GABA amount between 25 to 50 milligrams per 100 grams of weight, the CRISPR tomatoes had a wide range of increased accumulation, with the highest reaching around 250 milligrams per 100 grams of weight.
Compensating For Development
It was noted that while fruit size was generally not affected by this change, the amount of fruits produced and total growth and development of flowers was affected in some lines, though this varied significantly between different experimental groups, indicating that GABA accumulation and its impact on the plant can be larger or smaller depending on additional factors. There were several lines that were minimally impacted at the same time.
The researchers hypothesized that since the GABA accumulation was overexpressed in the plant tissues in general, including the leaves in particular, and not just the growing fruits, this affected flowering and overall yield. The specific biological impact observed is that high levels of GABA interfered with the elongation of certain cell types in the plant during development, so there was less growth overall. But they suggested that backcrossing into parental lines and the use of F1 hybrid traits would compensate for this and allow for the development of tomato cultivars with little to no negative growth effects and still producing high GABA fruits.
And that 2017 research has led to today, where we have the official release of the high GABA tomato. While the cultivar used is different from the specific kind used in the study, the same principles were utilized in its development (and likely will be used for many other varieties) and the altered genes appear to be stably inherited, so home growing and cultivation of the tomatoes can be done through seed packets. Hopefully this will lead to a greater adoption of these tomatoes in foods around the world and a global improvement in blood pressure issues for everyone.