Team:TU-Munich/HumanPractice/GMO

Example 1: Transgenic Crops

Example 2: Transgenic Moss for Phytoremediation

Assessment and Comparison

Figure 1: The Risk-Benefit balance symbolizes the cosideration of benefitial effects and potential risks that are an inheritent part of a specific technology.

Initiation

“What is the context between the three images in figure 1? “ Night blindness is a result of a so called vitamin-A- deficiency (VAD) .Directly in developing countries occur high rates of infection diseases and malnutrition which leads to low immunity. This effects night blindness which can cause complete blindness within few days . The golden rice project is a method of biofortification to combat the VAD with the aid of genetic engineering. The cleaning of the wastewater via the genetic engineered moss Physcomitrella patens in our project leads and aims also to improve the conditions directly in developing countries via genetic engineering.

The Golden Rice Project-humanitarian help with genetic engineering

“I was hungry and you didn’t feed me.” Matt. 25,42 “Let food be your medicine and medicine your food.” Hippocrates 460 BC "This rice could save a million kids a year" Time Magazine, 2000

Historical developing steps of the Golden Rice Project- from idea to reality

“Would it be possible to engineer the biochemical pathway of provitamin A into the rice endosperm to make the rice grain produce enough provitamin to reduce VAD of rice-dependent societies?”

To solve this question was the goal of the both inventors of the Golden Rice Technology: Prof. Ingo Potrykus (emeritus professor; ETH Zurich) and Prof. Peter Beyer (University of Freiburg; department Cell Biology). According to the World Health Organization (WHO) about 2, 3 million children die by reason of VAD and still 250000-500000 children per day go blind mainly in developing countries like Africa and Southeast Asia (see figure 2). To reduce the high rate of vitamin-A-deficiency and malnutrition Prof. Potrykus had the vision to improve the content of micronutrients in food crops. With the aid of the knowledge of the regulation of the terpenoid biosynthetic pathway in the model plant daffodil (Narcissus pseudonarcissus) from Prof. Beyer the plan of Prof. Potrykus completed. The following table (see table 1) should give you a short overview about the fundamental historical developing steps concerning the realization of the project idea. The role of the Chairman of the Humanitarian Board takes Potrykus. In addition to the progenitors and the financial support from the private sector many authorities and members of the Humanitarian Board lead to the benefit of this project in humanitarian genetic engineered help directly in developing countries (see figure 11). The Golden Rice Network is a community work with a strong trust and delivery which involves researchers, breeders, extensionists, and the small farmers. In general all technologies are patented. In total just 12 of the 70 patents were important for the developing countries which were found out by the help of the company Syngenta (before: AstraZeneca). The last has all worldwide commercial right through the treaty, in turn national and international research institutes was assured non-commercial use.So it was necessary to get just 12 licenses for the humanitarian project. It was contractually agreed that the GM-rice just used humanitarian not commercial i.e. the famers get and dispose the rice free and the industry had to abdicate money-disempowerment of license-frees. The GM-rice doesn’t need any herbicides and is expensive like non GM-rice. Also the farmers whose annual income is under $ 10,000 should use the GM rice without licensing costs.

Golden Rice – an impact of human health

“Why choose Beyer and Potrykus rice as the food crop for the genetically modification?”

Refer to table 1 Potrykus addresses with the difficult transformation and regeneration of monocots. The monocot rice is one of the basic staple food crops for half the humanity ( see figure 3) and a major energy source, representing the major carbohydrate and even protein source in SE Asia and Africa. The both inventors developed their project in South and Southeast Asia where 90 % (3 billion people) is grown and consumed rice. Concerning to figure 4 in some places more than half a kilo rice eats a person daily thus the energy intake per day comes from this food crop. Due to the brilliant source of calories the micronutrient content is very low. Another advantage is that the rice doesn’t contain provitamin A (more or less deficient of Vitamin A) which is necessary for humans to create vitamin A. So the lack of rice as a food can lead to VAD mostly affecting children and pregnant women. Rice includes high amount of substance which are needed basically (e.g. polished rice includes 20% of irons, folate, tiny amounts and zinc as the daily requirement).

The science of Golden Rice

“What is the golden on Golden Rice?” Compared to the non GM-rice the amount of β-carotene (provitamin A) in the Golden Rice is higher. Without genetic engineering methods the genetic variability of non GM-rice is too small to increase the content by conventional breeding techniques to β-carotene. In the β-carotene biosynthesis three enzymes are essential: phytoene desaturase (PDS), ζ-carotene desaturase (ZDS), ζ-carotene cis-trans Isomerase (Z-ISO) and carotene cis-trans isomerase (CRTISO) . The breakthrough of reconstitution of the carotenoid pathway in the grain and mdash in rice grains in 1999 (see table 1) was so essential for the advancement of the project. Naturally the rice plant produces in the leaves the enzymes PDS, ZDS and CRTISO .Compared to that the enzymes in the rice grain (endosperm of the rice) are down regulated. So the GM-rice in contrast to the Golden Rice is white due to the non building of the yellow -gold colored β-carotene in the rice grain. Figure 5 shows the engineered carotenoid pathway in grains of Golden Rice which leads to the golden color of the rice. In contrast to the non GM-rice they inset dissimilar genes to get β-carotene in the rice grain. The first used transgene encodes a plant phytoene synthase: NpPSY (from Narcissus pseudonarcissus) accelerates the reaction from GGPP (endogenously synthesised geranylgeranyl-diphosphate) to phytoene (colorless carotene with a triene chromophore) . To get phytoene to all-trans-Lycopin the three enzymes ZDS, PDS and CRTISO were replaced by bacterial phytoene desaturase (crt I) (from Pantoea ananatis; previously known as Erwinia uredovora) which leads to conjugation by adding four double bonds. NpLCY (from Narcissus pseudonarcissus) were used to react all-trans-Lycopin to β-carotene. From these dissimilar three genes (NpPSY, NpLCY and crt I) only two transgenes (crt I and PSY) are required to turn Golden Rice into a reality . Last based on the fact that the activity of rice LCYs is obviously not rate limiting, since lycopene doesn’t accumulate and the lycopene cyclase activities isn’t required to proceed down the pathway. In the GM-rice under the regulation of a tissue-specific Glutelinpromoter (gt1) instead of in the prototype used constitutive 35S promoter lead to an added production of β-carotene in the rice grain. The last works because of the exclusive effect of the Glutelinpromoter in the rice endosperm. Consequently the Golden Rice is yellow-golden colored because of the activity of intrinsic rice cyclases.

“What is the intension of producing new Golden Rice generations?”

In further experiments they produced Golden Rice 1 (GR1) and Golden Rice 2 (GR2)(see figure 6). One difference to the first developed Golden Rice prototype is that the used transgenes (NpPSY,crt I) are regulated by the tissue-specific Glutelinpromoter (gt1) instead of in the prototype used constitutive 35S promoter. The last works because of the exclusive effect of the Glutelinpromoter in the rice endosperm. Experiments with PSY genes which are the more active compared to crt I from different sources identified the maize and rice genes as the most efficient in rice grains . Golden Rice 2 contains just ZmPSY and crt I in contrast to Golden Rice 1 (crt I and NpPSY). Another phytoene synthase (psy) from another plant were used because of the fact that the Phytoensynthase of the plant corn (zea mays) had a better achievement as the plant daffodil ( Narcissus pseudonarcissus). Another reason for the development of new Golden Rice generations was the resulting amount of β-carotene levels. Table 2 shows a list of the resulting β-carotene levels by using the different rice generations. To combat VAD in South Asia it is necessary to provide adequate amounts of provitamin A in children's diets. Just the amount of 31 µg/g of β-carotene covers the daily provitamin A requirements of the target population in the absence of a more varied diet to maintain appropriate levels of vitamin A in the body. This underlines the need of GR2 and difference to the other generations. “Why is Golden Rice undoubtedly an excellent source of the vitamin A ?” In addition to other factors the vitamin A status is the most important which influences the absorption and utilization of provitamin A carotenoids. The carotenoid content isn’t alone sufficient to list how much takes up by the body because of the affect of the other factors to the bioavailability of carotenoids, carotenoid species, concentration, the food matrix, dietary fat, and also the health status of the person. So to facilitate provitamin A absorption dietary fat is necessary. In contrast to that eating of Golden Rice leads to maintain appropriate vitamin A blood levels and absorbtion of sufficient provitamin A from the diets without needed oil. So the intestinal β-carotene uptake gets facilitated by the simple starchy food matrix of the endosperm and the fat content of the rice grain.

Vitamin A deficiency

Malnutrition is a vehement problem which demands the death of 12 million children worldwide. The damage of micronutrient deficiencies in the world is caused by the low dietary intake of iron, iodine, zinc and vitamin A. Concerning to the WHO (2012) 2.7 children can save from death via providing those with vitamin A. Directly preschool children and pregnant woman of the poor population are affected by the VAD in Southeast Asia. Sadly rice is a poor source of many essential micronutrients (see figure 7) . So micronutrient malnutrition causes by a rice based diet in Africa and SE Asia. “Why is Golden Rice a blessing to combat the VAD?” Next to traditional interventions like distribution of retinol pills, accumulation of food with micro nutritive substances, diet diversification, disease control and disaster relief is this biofortification with genetic engineering another strategy to improve the malnutrition situation and combat the VAD. Naturally the plant rice can convert provitamin A (β-carotene) into vitamin A (retinol derivates) just in the leaves. The dependence on rice as the predominate staple food crop can reduce to use genetic engineering. The Golden Rice project is one example of producing GM rice to convert the provitamin A into vitamin A in the grain. Even if this project leads to an effort concerning the malnutrition in Southeast Asia the eradication of VAD and malnutrition can’t happened without the solving of underlying political, economic and cultural issues, the dispersion of supplementation programs and the achievement of financial support worldwide. The medical consequences of VAD:

• impaired vision
• irreversible blindness
• Night blindness
• increased risk of atherosclerotic heart disease
• fertility disorder
• impaired epithelial integrity
• susceptibility to infection
• dryness of the skin
• reduced immune response
• defective haemopoiesis leads to reduced capacity to transport oxygen in the blood
• disturbed skeletal growth
• etc.

“Which Strategies are available to combat Vitamin-A-deficiency?” In addition to the biofortification strategy (e.g. Golden Rice) there are also classic intervention approaches of supplementation or fortification. In general no single intervention solves micronutrient malnutrition i.e. all different strategies are important. The following apron shows you a few strategies for combat VAD within the limitations to each approach.

1.Biofortification: “biofortified” means genotypes which deliver increased levels of essential vitamins and minerals by use of plant breeding of micronutrient content of crops. In contrast to the supplementation programs biofortifiction leads to providing a daily, sustainable supply of dietary nutrients. Another advantage is the efficient access to the technology in form of seeds. In comparison to the supplemetation programs in India normally 2,700 $ against 4,300 $ are necessary to save each life concerning to an article in ZM Science . Present ambitions are aiming at adding other micronutrients to the rice grain (e.g. iron, zinc, high-quality protein, and vitamin E) after introgressing the provitamin A-producing trait into local varieties has happened sufficient. 2.Supplementation Programs: Concerning to the cost analysis of the national supplementation programs in Ghana, Zambia and Nepal in 2004 it shows clear the disadvantage “cost” of the supplementation programs. Twice a year capsules of vitamin A in solution were distributed to six months to five year aged children. Total 1,14 $ had to paid for annual cost per child doses. Other limitations are the opportunities for frequent contact, the support of daily doses to the population especially children and pregnant women and the logistic problems like need for training, medical staff and distribution infrastructure.

Another strategy is the accumulation of food via manufacturing or packaging with micronutrients which is just efficient by consuming the processed industrial products. The diversification of nutrition i.e. measures like education about consuming more Vitamin A containing food. The problem of this strategy is the regional and seasonal availability of Vitamin A containing food.

Introduction

Legal regulations in the European Union

Legal Regulation World Wide

Actual Situation in Germany

http://www.ncbi.nlm.nih.gov/pubmed/6327079 Edens et al., 1984

1. http://www.ncbi.nlm.nih.gov/pubmed/6327079 Edens et al., 1984 Edens, L., Bom, I., Ledeboer, A. M., Maat, J., Toonen, M. Y., Visser, C., and Verrips, C. T. (1984). Synthesis and processing of the plant protein thaumatin in yeast. Cell, 37(2):629–33.