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On path to making cereals that consume nitrogen from air

Solving “nitrogen problem” by transferring nitrogenase genes to plants would generate crops much less dependent on chemical nitrogen fertilizers.

Some bacteria carry an enzyme called nitrogenase that enables them to grow by incorporating nitrogen from the air into biomass. Transferring nitrogenase to cereals would keep crop production high while reducing nitrogen fertilization, according to information from the Universidad Politecnica de Madrid in Spain.

Although nitrogen gas is the major component of the atmosphere, it is biologically inactive for plants, fungi and animals. Only some bacteria, called diazotrophs or nitrogen eaters, are able to convert (fix) nitrogen gas into ammonia, which they and other living organisms then use to grow biomass. Diazotrophs are, therefore, the primary producers of nitrogen in all ecosystems.

Agricultural systems in developed countries are largely based on cereal crops, which provide about 60% of the energy in people's diets, the university said. Nitrogen and water availability are the most important factors limiting cereal crop productivity. Unlike legumes, cereals are unable to form symbioses with diazotrophs and their production is tied to the application of nitrogen fertilizers.

During the last 100 years, cereal crop yields have increased via the addition of chemically synthesized nitrogen fertilizers — at a cost exceeding $100,000 million per year. Pervasive use of nitrogen fertilizers in developed countries has degraded the environment to a point that, at times, is incompatible with animal life in marine ecosystems, the university said. In contrast, the high cost and limited availability of chemical fertilizers prevent its use by poor farmers, causing poverty and hunger derived from low crop yields.

Diazotrophs use a two-component enzyme called nitrogenase to fix nitrogen. Because plants lack nitrogenase, they cannot “eat” nitrogen gas.

The laboratory of professor Luis Rubio at the Universidad Politecnica de Madrid is trying to solve the so-called “nitrogen problem” by transferring nitrogenase genes (nif) to plants. This would generate diazotrophic plants that are much less dependent on chemical nitrogen fertilizers.

This is considered one of the most difficult challenges in plant biotechnology. Because nitrogenase is destroyed by oxygen gas, it has long been assumed that it would not work in cells from plants, animals or fungi, all of which require oxygen to produce energy. Such incompatibility would be exacerbated in plant cells producing oxygen as result of photosynthesis.

However, a recent study from Rubio's laboratory demonstrated that the most oxygen-sensitive component of nitrogenase functions in yeast (a fungi used as model to expedite outputs) as long as it is kept inside mitochondria, an intracellular organelle responsible for oxygenic respiration.

Working together, Rubio's laboratory and the laboratory of professor Christopher Voigt, a leading synthetic biologist at the Massachusetts Institute of Technology, have now taken a second step in assembling nitrogenase by transferring nine nif genes to the genome of yeast and expressing them at controlled levels. This study proved the correct formation of the other nitrogenase component in mitochondria, an essential step of nitrogenase assembly. The study also showed that nitrogenase was not fully functional and that further bioengineering is needed to obtain active mitochondrial nitrogenase.

The research performed in Rubio's laboratory at the Centro de Biotecnología y Genómica de Plantas of Universidad Politécnica de Madrid is funded by the Bill & Melinda Gates Foundation and aims to help smallholder farmers in sub-Saharan Africa. Although much bioengineering remains to be done, research carried out at the lab will hopefully contribute to reducing nitrogen fertilization in developed countries and to increase cereal production for farmers in developing countries of Africa and Asia.

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