Taste the Sun: Gene Editing Produces Vitamin D Enhanced Tomatoes

Date:06-23-2023

The applications of CRISPR gene editing in agricultural biotech have boomed in recent years, from improving crop yield to maximizing food nutrition. One of our favorite red fruits, the tomato, is no stranger to the CRISPR toolbox. In 2021, genome edited tomatoes enriched in γ-aminobutyric acid (GABA) were commercially launched in Japan by Sanatech Seed.

In a new report published in GEN Biotechnology (the sister journal to GEN), researchers from Seoul National University (SNU) in South Korea have applied CRISPR gene editing to produce tomato fruits with enhanced levels of provitamin D3 (ProVitD3).
In humans, ProVitD3 is a precursor for the synthesis of biologically active vitamin D3 and serves as a protective agent against ultraviolet radiation on the skin. Circulatory vitamin D often falls below desirable levels, particularly among individuals with limited exposure to sunlight. While dietary supplements help address this deficiency, natural vitamin D sources are restricted to a few animal-derived sources, such as fish, egg yolks, and beef liver, as fruits and vegetables have historically shown limited capacity for ProVitD3 production.

In the new study—featured on the cover of the June issue of GEN Biotechnology—Sunghwa Choe, PhD, and his colleagues at SNU used CRISPR gene editing in tomatoes to induce a loss of function in one of two DWARF5 (DWF5) genes, a homologue of the human gene (DHCR7) responsible for converting ProVitD3 to cholesterol. The plant homologue of DHCR7 was initially identified in Arabidopsis thaliana as DWF5.

Given a high sequence identity of more than 83%, the Korean team hypothesized that the two tomato DWF5 genes, SIDWF5A and SIDWF5B, function redundantly in their biosynthetic pathways. They chose to edit SIDWF5A after spatial expression patterns showed that transcript levels of SIDWF5A were 2–5 times higher than those of SIDWF5B, especially in green and red fruits.
Remarkably, the new gene-edited tomatoes showed accumulated ProVitD3 levels as high as 6 μg/g dry weight (DW) in red fruits, while maintaining morphological similarity to wildtype tomatoes. (By contrast, Arabidopsis dwf5 mutants display dwarfism.) As the daily recommended human intake of vitamin D is 20 μg, the authors state that consuming a single ripe fresh tomato weighing 150 g (equivalent to 15 g DW) has the potential to significantly alleviate vitamin D deficiencies worldwide.

″With this work, Choi et al. show a second proof that tomato fruits can be biofortified with elevated levels of ProVitD3, which could provide a plant-based source of vitamin D if plants with this trait are brought to market,″ comments Aaron Hummel, PhD, chief technology officer at Pairwise, a North Carolina-based food tech company applying CRISPR to develop ‘Conscious Greens’ salad mixes that aim to be fresher, tastier, and easier to access.

″This is another important step in a groundswell of nutritional, quality, and other beneficial improvements that are being made in fresh fruit and vegetable crops by a wide range of public and private institutions and companies. I’m pleased to see the progress in this space that is made for the benefit of the end consumer,″ Hummel said.

While these ProVitD3-enhanced tomatoes are promising, the authors note that further research is needed to quantify and compare the levels of ProVitD3 in genome-edited tomatoes with other natural sources of vitamin D3. Notably, stability and bioavailability studies are needed to evaluate the impact of cooking, processing, and storage on the retention of ProVitD3 in addition to feeding studies and clinical trials to determine the bioactivity and physiological effects of consuming ProVitD3-enriched tomatoes.

The authors also note that some of the genome-edited lines display a lower number of fruits, although this variability could be mitigated by introducing the ProVitD3 trait into genetically fixed inbred lines.

CRISPRing more tomatoes

In addition to following in the footsteps of Japan’s GABA tomatoes, this GEN Biotechnology study expands upon the related results published in Nature Plants in 2022 by Cathie Martin, PhD, and colleagues from the John Innes Centre in the U.K., which demonstrated the accumulation of ProVitD3 in stems and leaves through knockout of SIDWF5B. By editing the sister gene, SIDWF5A, the Korean group produced higher levels of ProVitD3 accumulation, specifically in fruit tissues.

Professor Choe, who is also the CEO of a company called G+FLAS Life Sciences, says that researchers are actively exploring other modifications of tomatoes to enhance both yield and nutrition using CRISPR. A recent review article lists leaf, stem, flower, male sterility, and fruit development, as a few target tomato traits.

In addition, researchers are enhancing biotic stress resistance against pathogens, such as tomato yellow leaf curl virus, powdery mildew, and late blight, and improving abiotic stress tolerance, such as resistance to heat, cold, drought, salinity, and enhancing carbon-nitrogen metabolism. Herbicide resistance is another area of interest.

And the GEN Biotechnology authors are not stopping at tomatoes. Research is underway to introduce this trait to other crops that contain at least two copies of the DWF5 genes, including pepper and paprika crops.

To the market?

The burning question remains: What are the regulatory and scale-up hurdles that impact whether these tomatoes head to the market? Choe states that in many countries, there is a growing consideration that CRISPR genome-edited crops may be exempt from conventional GMO regulations.

″Specifically, plants edited using site-directed nuclease 1 technology are often deemed indistinguishable from naturally occurring spontaneous mutations and therefore exempt from regulatory oversight,″ said Choe. ″South Korea has also initiated discussions on this matter, and the Ministry of Trade, Industry and Energy (MOTIE) has submitted a revision to the Living Modified Organisms Act to the National Assembly of the Republic of Korea.″

Moreover, the governmental approval process for the GABA tomatoes in Japan has served as a guide for other countries considering the commercialization of gene-edited crops.

″Our company takes into account the Japanese GABA cases to facilitate discussions with the general public and key government agencies, including MOTIE, the Ministry of Agriculture, Food and Rural Affairs (MAFRA), the Ministry of Food and Drug Safety (MFDS), the Ministry of Maritime Affairs and Fisheries (MOMAF), and the Ministry of Environment (MOE),″ Choe told GEN. ″By drawing on the experience and regulations surrounding GABA tomatoes, we aim to navigate the approval path for future gene-edited tomatoes.″

As for scale-up, Choe says his team is actively preparing commercial seeds while awaiting the resolution of regulatory challenges in Korea. Simultaneously, the researchers are exploring collaborations with global seed companies. The seeds and technologies are also readily available for out-licensing. Choe’s goal is to release these gene-edited fruits for sale as early as 2026, once regulatory hurdles are resolved in Korea and elsewhere.

When asked about the public perception in Korea to CRISPRed fruit, Choe remained encouraged.

″While there are strong voices from anti-GMO activists in Korea, it is important to note that there is also a growing demand for functional and premium-grade tomatoes among the general public,″ Choe said. ″Korean consumers are becoming more aware of the fact that Korea is the only country in Northeast Asia where the regulatory status of CRISPR-edited crop plants is not clearly defined as either Living Modified Organism (LMO) or Non-LMO.″

Choe says that lawmakers in Korea are actively engaged in discussions regarding this issue, indicating a willingness to explore and potentially relax regulations surrounding CRISPR-edited crops in the coming years. At the same time, Choe emphasizes the importance of ongoing public engagement, education, and transparent communication to ensure a well-informed and inclusive decision-making process.