West Lafayette, Indiana
June 17, 2002

Forget the attack of the killer tomato, this is the attack of the healthy tomato: A team of scientists has developed a tomato that contains as much as three and a half time more of the cancer-fighting antioxidant lycopene.

It turns out that the antioxidant-rich tomato was a happy accident.

Scientists at Purdue University and the U.S. Department of Agriculture's  Agricultural Research Service were working to develop tomatoes for food processing that were of higher quality and would ripen later.

They accomplished that, but in the process they discovered that the new tomatoes also had significantly more of the antioxidant than conventional tomatoes.

"We were quite pleasantly surprised to find the increase in lycopene," says Avtar Handa (pronounced "Honda"), professor of horticulture at Purdue.

Although increasing the nutritional value of foods is the goal of so-called second-generation biotechnology products, there have been few success stories.

"This is one of the first examples of increasing the nutritional value of food through biotechnology," Handa says. "In fact, it may be the first example of using biotechnology to increase the nutritional value of a fruit."

Co-discoverer Autar Mattoo, who heads the USDA Vegetable Laboratory, says the increase in lycopene occurred naturally in the genetically modified tomatoes. "The pattern for the accumulation was the same as in the control tomatoes," he says. "The lycopene levels increased two to 3.5 times compared to the non-engineered tomatoes."

The research was announced this week in the June issue of Nature Biotechnology.

A separate article on the research in Nature Biotechnology noted, "The findings Š remind us that in the 'rational' and quantitatively driven post-genomic era, serendipity still has a large part to play."

A U.S. patent application has been filed on behalf of the joint owners USDA and the Purdue Research Foundation. The Consortium for Plant Biotechnology Research Inc., a USDA funded program, funded the research.

Lycopene is a pigment that gives tomatoes their characteristic red color. It is one of hundreds of carotenoids that color fruits and vegetables red, orange or yellow. Of these pigments, the most familiar is the beta-carotene, which is found in carrots.

In the body these pigments capture electrically charged oxygen molecules that can damage tissue. Because of this they are called antioxidants.

Lycopene has been the focus of much attention since 1995, when a six-year study of nearly 48,000 men by Harvard University found that men who ate at least 10 servings of foods per week containing tomato sauce or tomatoes were 45 percent less likely to develop prostate cancer. The study also found that those who ate four to seven servings per week were 20 percent less likely to develop the cancer.

That research was published in the Journal of the National Cancer Institute.

Subsequent research has found that lycopene also reduces the amount of oxidized low-density lipoprotein - the so-called bad cholesterol - and therefore may reduce the risk of heart disease.

As an antioxidant, lycopene is able to capture twice as many oxygen ions in the body as is beta-carotene.

"This characteristic may be responsible for lycopene's ability to mitigate epithelial cancers, such as breast cancer and prostate cancer, and for its ability to mitigate coronary artery disease," Mattoo says.

Despite the apparent benefits, it's been difficult to increase the amount of lycopene in the diet, says Randy Woodson, director of Agricultural Research Programs at Purdue.

Studies have found that taking purified antioxidants as a dietary supplement doesn't work. In fact, one study found that giving beta-carotene to smokers actually increased their chances of developing cancer.

"When you just take lycopene as a drug it doesn't have the same effect," Woodson says. "There is still a lot of biology to understand before we know why phytonutrients in food are so much more effective than if they are given as supplements."

Another wrinkle is that when it comes to lycopene in tomatoes, cooked tomato sauces are more effective than raw tomatoes.

This may be because cooking breaks the cell walls of the tomato, releasing more of the lycopene. Or it may be that cooking oil allows the lycopene to move more easily into the body.

To develop the lycopene-rich tomato, the researchers inserted a gene, derived from yeast, fused to a promoter gene into tomato plants. The promoter gene helps turn on the yeast gene in the tomato.

"The promoter gene is like a ZIP code that tells the yeast gene when and where to turn on in tomato," Handa says. "For high-lycopene tomatoes we used a promoter that targeted expression of the introduced gene in fruits only."

The yeast gene itself produces an enzyme that affected the production of growth substances in the plants called polyamines, which are known to help prevent cell death.

In plant cells, polyamines help build new, beneficial compounds.
 
"They may stabilize membrane networks that involve longevity of physical structures in the cells called chromoplasts," Mattoo says. "Because lycopene accumulates in chromoplasts in the tomato fruit cells, in this case the polyamines seem to have a positive effect."

The polyamines share a precursor with a plant hormone called ethylene that causes ripening in many fruits.

The researchers thought that because ripening was delayed there must have been a decrease in ethylene, but found the opposite was true.

"That's not how we started out thinking, but that's why we do experiments," Mattoo says. "Now we know the change ‹ i.e., allowing the accumulation of polyamines in the fruit ‹ doesn't necessarily affect ethylene production, but ethylene action. We think the polyamines has changed the ethylene receptors on the cell membranes, but we are looking into that."

Handa says the technique used in this research might also be used to increase the amount of other antioxidants in foods.

"We are excited about this approach, not only because it results in an increase in lycopene in tomato, but because we think this approach could be used to increase the phytonutrient content of other fruits and vegetables," he says.

Writer: Steve Tally, (765) 494-9809; tally@purdue.edu

Sources:
Avtar Handa, (765) 494-1339, handa@hort.purdue.edu
Autar Mattoo, (301) 504-6622, mattooa@ba.ars.usda.gov
Randy Woodson, (765) 494-8362, woodson@purdue.edu

Related Web sites:

Nature Biotechnology: http://www.nature.com/nbt/

Nature Biotechnology press release on this research:
http://www.nature.com/nbt/press_release/nbt0602.html

2000 USDA-ARS article about the tomato research:
http://www.ars.usda.gov/is/AR/archive/sep00/tomato0900.htm

Handa's Web page: http://www.hort.purdue.edu/hort/people/faculty/handa.html

Mattoo's Web page: http://www.barc.usda.gov/psi/vl/mattoo.htm

ABSTRACT

Title Engineered polyamine accumulation in tomato enhances phytonutrient content, juice quality and vine life

Roshni A. Mehta1, Tatiana Cassol1, Ning Li2, Nasreen Ali1, Avtar K. Handa3, and Autar K. Mattoo1

1USDA-ARS Henry A. Wallace Beltsville Agricultural Research Center, Beltsville, MD; 2The Hong Kong University of Science and Technology, Hong Kong SAR, China; 3Purdue University, West Lafayette, IN

Polyamines, ubiquitous organic aliphatic cations, have been implicated in a myriad of physiological and developmental processes in many organisms, but their in vivo functions remain to be determined. We expressed a yeast S-adenosylmethionine decarboxylase gene(ySAMdc: Spe2) fused with a ripening-inducible E8 promoter to specifically increase levels of the polyamines spermidine and spermine in tomato fruit during ripening. Independent transgenic plants and their segregating lines were evaluated
after cultivation in the greenhouse and in the field for five successive generations. The enhanced expression of the ySAMdc gene resulted in increased conversion of putrescine into higher polyamines and thus to ripening-specific accumulation of spermidine and spermine. This led to an increase in lycopene, prolonged vine life, and enhanced fruit juice quality. Lycopene levels in cultivated tomatoes are generally low, and increasing them in the fruit enhances its nutrient value. Furthermore, the rates of
ethylene production in the transgenic tomato fruit were consistently higher than those in the nontransgenic control of fruit. These data show that polyamine and ethylene biosynthesis pathways can act simultaneously in ripening tomato fruit. Taken together, these results provide the first
direct evidence for a physiological role of polyamines and demonstrate an approach to improving nutritional quality, juice quality, and the vine life of tomato fruit.