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Unit 13: Genetically Modified Organisms
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Gary H. Toenniessen, PhD.

Gary Toenniessen was one of the original architects of The Rockefeller Foundation Rice Biotechnology Research Program initiated in 1984. He is currently the director of Food Security at the foundation. Since 1985, he has had responsibility for the development and implementation of the Foundation's International Program on Rice Biotechnology, designed to bring the benefits of biotechnology to poor rice producers and consumers in developing countries.


How did you get involved with the creation of Golden Rice?

The Rockefeller Foundation exists for the benefit of mankind, and was set up by John D. Rockefeller in 1913, and has worked for many years to try to improve the living conditions for people in developing countries.

Since the 1940's, that has included work on agriculture, and in the 1950's and '60's the Foundation made major investments in the genetic improvement of the staple food crops in Latin America, Asia, and Africa. Rice is the most important food crop in the developing world, particularly in Asia, so a lot of that work went into rice. The Foundation helped to establish the International Rice Research Institute in the Philippines, helped to build the national rice breeding programs in India, Thailand, Indonesia, Pakistan, and many other countries of Asia, and that was a very successful program-it led to the Green Revolution that occurred in Asia.

What is the importance of rice as a crop?

Rice is extremely important in Asia. If you take a country like Bangladesh, 80% of the calories that are consumed by the people are derived from rice. There's nothing equivalent to that in the United States. Our most important crop is wheat, and only maybe 20% of the calories that we consume are derived from wheat.

And the poor people are more dependent on rice, so there are people in Asia who only eat rice, and they're lucky if they get something to accompany that rice.

What is rice lacking in terms of nutritive components?

Rice is a great source of calories. It's also a good source of protein. The protein in rice is a fairly balanced protein. It contains all of the essential amino acids. What rice really lacks are vitamins and minerals, the kinds of micronutrients that one gets from vegetables or from meat. So vitamin A is a key vitamin, but most of the other vitamins are lacking from rice as well. It also has low levels of iron, zinc, iodine, the other nutrients that are essential for human nutrition.

Why are vitamins lacking in rice?

First of all, you need to realize that rice when it's consumed is polished. The outer layers of rice are removed in the process of storing rice. If they're not removed, the rice will turn rancid and spoil.

So the rice grain itself, the part that is consumed by most of the people in Asia contains no Vitamin A at all, and none of the precursors to Vitamin A, which would be beta-carotene and other carotenes.

Who is affected by this lack of Vitamin A?

Children tend to be most severely affected by the Vitamin A deficiency-particularly children during the weaning period. When the children are taken off of breast milk, they're fed primarily a rice-based diet.

Are there problems with long-term deficiency?

Well, there is now a lot of evidence that Vitamin A deficiency causes numerous problems, most of them are related to malfunctioning of mucous membranes throughout the body. The most obvious result of this is night blindness. But in addition to that, there is good evidence now that mucous membranes throughout other parts of the body have been weakened, which makes the body more susceptible to other diseases, diseases such as measles.

So, in developing countries, a lot of children still die of diseases such as measles and there's good evidence now that Vitamin A deficiency makes these children more susceptible to those diseases because their immune system is weakened, and the membranes that protect them from infection in their lungs and their respiratory tract and their digestive tract are weakened.

Why is there interest in producing beta-carotene in rice?

We were interested in rice improvement, so we were interested in many traits. In fact, we had identified a list of some 25 traits that we were encouraging researchers to investigate. One of those was beta-carotene production in the rice grain. That is because the rice scientists, who for years have been working to improve rice, recognized its nutrient deficiencies and had been searching for a natural variant of rice that did produce beta-carotene.

They thought such a variant might exist because other plants have produced such variants. Yellow corn, for example, is a mutant. Wild type corn or natural corn is white. Carrots are the same way. Wild type carrots have white roots. At some point in time, man selected an orange-colored carrot, either because he found it attractive, or maybe he or she recognized that it had some nutritional value associated with it.

So these are variants where beta-carotene is produced in a tissue where it really doesn't have its traditional function. Beta-carotene is a photosynthetic pigment. It occurs in all green plants in the green tissues of those plants. It's there to protect the chlorophyll.

If beta-carotene and other pigments like beta-carotene do not exist in the green tissue, the chlorophyll becomes bleached by the sun, so it's an essential component of green tissues, including rice. So rice already produces beta-carotene in the leaves, in the stem, and the other green tissues. It doesn't have its traditional function in non-green tissues, so under normal conditions it's not produced.

Were there skeptics who thought it was impossible?

The rice breeders had been looking for this to occur naturally. They'd been looking for a strain of rice that produced beta-carotene, and there actually are some rices that are colored, but it turned out that none of them were producing beta-carotene.

So what that meant was that you couldn't use conventional breeding. There was no natural genetic source of a rice, or even a wild relative, that you might be able to cross into rice that produced beta-carotene. That meant that we would need to use genetic engineering approaches, where you actually added genes to the plant in order to produce the beta-carotene, or somehow figure out how to turn on the genes that were already in the rice plant in the grain tissue where they were not normally turned on.

Both of those were going to be extremely difficult and complex research projects, and there were some advisors who told us that they were too difficult, that the Foundation would be better off spending its money on traits that were easier to manipulate using these techniques where all you had to do was add one gene.

How was the idea formulated of how to get the rice to produce beta-carotene?

We put together a workshop in which we invited about 20 people who were knowledgeable about the importance of Vitamin A in rice, and two or three nutritionists in that area. Also, a biochemist who knew about the beta-carotene biosynthetic pathway, Peter Beyer.

By the end of the workshop we actually came up with two strategies that looked like they were worth pursuing in order to try to achieve that goal, and we wound up funding both of those strategies. One of those was to introduce all of the enzymes that would be required to convert the precursor that we knew existed in rice to beta-carotene and that would require four enzymatic steps. That was the research project that Professor Ingo Potrykus and Peter Beyer submitted to us.

So they thought it was a feasible process?

They wouldn't have done it if they didn't think they could. We wouldn't have funded it if we didn't think there was a chance. I think all of us recognized that it was a high-risk project. Nobody had introduced four genes into a plant and gotten them to function in a sequential way that allowed a whole biosynthetic pathway to be introduced previously, so it was cutting edge research, but they were some of the best in the field and we felt it was worth taking a chance.

What was the first discovery about the pathway?

One was that the precursor did exist in the rice grain; the precursor is called geranyl geranyl diphosphate. The research that we supported shows that it did exist at levels that would be nutritionally significant if it were converted to beta-carotene. That preliminary research also showed that phytoene, which was the first compound in the carotenoid pathway was absent from rice endosperm. So, the first step was missing but the materials in order to achieve that first step were present.

Why did the researchers look to daffodils?

Peter Beyer's area of research interest is the carotenoid biosynthetic pathway. He had chosen daffodils to study because they're a good model plant. They produce beta-carotene in the flower. The flower is the same as grain. It's not a photosynthetic tissue, and therefore there are a variety of mutants of daffodil, so he could look at daffodil plants that did or did not produce beta-carotene, and that did or did not produce various other carotenoid compounds in this pathway. So it was a good plant to study the biochemistry of the carotenoid pathway.

What was the next step?

The first milestone that we were looking for was to see whether or not we could produce phytoene, the first 40-carbon compound on the pathway.

So, what Professor Potrykus wanted to achieve was for the introduced gene to be only expressed in the grain, because he was concerned if it were expressed in other tissues that it might have some deleterious affect in those other tissues.

What convinced Rockefeller Foundation that this might actually work?

When Potrykus added the phytoene synthase from the daffodil that Peter Beyer had provided him with, the rice plants produced phytoene. That was very encouraging, because that was the first step in the pathway. That showed that the precursors could be converted by this introduced gene into a carotenoid compound.

Following that, there were a number of disappointments. When they tried to introduce some of the other genes, one in particular was not able to carry out the desired step in the pathway, or at least the daffodil gene was not able to carry out that particular step. And so there were discouragements as well as encouraging steps along the way.

What was the breakthrough?

There were two breakthroughs. One was to try a different gene, rather than the daffodil gene, and one of the scientists was studying a similar pathway to this that occurs in bacteria. There are some bacteria that also produce carotenoid compounds, and he had isolated a gene from a bacterium called Erwinia, and that particular gene produced an enzyme that carried out two steps in this pathway.

Professor Potrykus had originally envisioned that what he would do was introduce these genes independently into rice plants and then cross the plants as occurs naturally. And he had a Chinese postdoctoral scientist working with him who actually had a limited period of time in which to help out on this research. So in addition to following that particular strategy, his Chinese postdoctoral scientist said, well, I'm going to try to add all of these genes at one time to the rice plants, and it turned out that was the was the breakthrough that they needed. This technique of introducing all of the genes together at one time led to the first of the rice plants that produced grain that had the desired golden color. Then Peter Beyer was able to show that that golden color was in fact beta-carotene.

How long did it take to create the pathway?

The workshop was held in 1993. I believe this breakthrough was accomplished in about 1999, so this was a six-year period of time. What they were actually able to do was introduce two genes from daffodil, each with a promoter and a leader sequence, and this one bacterial gene and in combination, those three genes were able to produce the enzymes to carry out the four enzymatic steps to convert the precursors to beta-carotene.

How does Golden rice differ from other commercial GMO's?

The objective was to produce a product that would benefit consumers. This is not a product that's going to benefit the farmer. The farmer isn't going to produce more rice, or cheaper rice, and it's not going to be drought tolerant. So it's one of the few examples of a research project that had as its principal objective the production of an end product that was aimed at benefiting the consumer and particularly, in this case, improving the nutritional value of the rice.

It's also unique in that it did involve modifying a pathway. Most of these other genetically engineered crops introduce a gene which produces a protein in which that protein itself is the desired end product. Well, in this case the genes that were introduced produced enzymes, and those enzymes carried out enzymatic steps that led to the desired end product, which was beta-carotene.

How did the project work?

Well, the science itself worked because of the collaboration of the main scientists, but also the willingness of all of these other scientists to share materials that they had developed, the promoter, the leader sequence, the bacterial gene, all of that was readily shared with Professor Potrykus.

It really wasn't until the product was available-the golden rice was available-that the issues related to intellectual property rights and bio-safety came into play-and it's at that time that the private sector, the companies, became involved and were able to make important contributions to moving the project toward an application and actual use of this very exciting product.

What is the goal now for Golden rice?

Well, first of all, you need to recognize that the first product was really just a proof of concept, and Professor Potrykus and Peter Beyer have continued their research in order to produce a product that really can be distributed to farmers. So what they've tried to do is remove some of the materials that were introduced along with these genes for research purposes, for example, an antibiotic resistance gene existed in the original material. They've now eliminated that.

Also, the newest version of golden rice has only two genes added to it. It still functions. It still produces beta-carotene. So, from a research standpoint they are producing a better product. In the Philippines and in a couple of other Asian countries, they are field-testing the materials that are being produced in Switzerland.

What kind of testing is being done in local countries?

In the Philippines and in India there are collaborating scientists who are now, first of all, field testing these materials, making sure that when it is grown that it still has all of the other desirable characteristics that you would want in a rice plant, but they are also crossing the golden rice with the varieties that are most important in the regions of their countries where Vitamin A deficiency is also most important.

What will be the cost to farmers for the rice seed?

It won't cost them anything in addition to what they already pay for rice seed. That varies. In some countries, the government supports programs that provide farmers with rice seed, new varieties of rice seed, so if the government is providing farmers with free seed then they will get the golden rice for free.

How do you answer to the skeptics?

One of the criticisms you most often hear is that there's not enough beta-carotene in Golden rice to make a difference. When you hear that criticism, people tell you the average daily requirements of a child for pro-Vitamin A.

What you need to realize, however, is that these people are deficient in Vitamin A. Vitamin A is not totally absent from their diet. So these children lack maybe 10%-20%, probably at the most 50% of the recommended daily requirement.

So if golden rice provides say only 10% of the recommended daily allotment of pro-Vitamin A, that may be enough to solve the deficiency of a significant segment of the population that is Vitamin A deficient. It's not going to be the only source of Vitamin A, and no one ever expected it to be the only source. But it clearly can make an important contribution.

We still have 200 million people who are Vitamin A deficient. This is another strategy that can make a contribution toward addressing that problem, and it can do so for people that are very difficult to reach using other mechanisms. Fortification and supplementation are very effective in urban areas. They're not very effective in rural areas where people consume the rice they grow. If you're going to fortify food, it has to go through a processing process.

If you buy rice in the United States, you can look on the back of the rice you buy and it's fortified. In fact, it's fortified by law in the United States. Why shouldn't children in developing countries be able to have the rice they grow be fortified the same way that our rice is? This is a way of making a contribution toward assuring that that could happen.

Is there an effort to produce more beta-carotene in the rice grain?

Part of the research that Professor Potrykus and Peter Beyer are currently doing in order to produce a product that can be commercialized-that can be distributed to farmers-is also to try to increase the amount of beta-carotene that can be produced. They've been successful to a degree, but not to the degree that they would like to be, so there is research continuing to see how that might be done.

Would you like to add anything else?

I think that the key thing is that this is not a silver bullet. Nobody is saying that this is the solution to the Vitamin A deficiency problem. To solve Vitamin A deficiency, the world, and particularly the countries where this is a serious problem, is going to have to implement a variety of strategies. They're going to have to work on providing a more diversified diet. They're going to have to work on supplementation and fortification.

I might also add that if it works in rice, which it does, this opens up the opportunity of doing similar research with other crops and improving their nutritional value as well, and not just with Vitamin A, but with other vitamins and with micronutrients. So it sets the stage for a whole new strategy for dealing with human nutrition problems in developing countries.


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