Drought Tolerance Association Mapping

Drought is a potential major constraint to maize production in all areas where it is grown, but it is a greater problem for the rural poor of developing countries. According to the Colombia University Center for Hazards and Risks, drought caused as many deaths in 1980-2000 as all other natural disasters combined. Drought is second only to poor soil fertility in reducing yield in the developing world, leading to a 15% overall reduction in grain yield in these countries, and in a bad year can be the major constraint on yield in developed countries as well. Global warming, deforestation, and urbanization will all increase the severity and frequency of drought in the future, leading to a possible decrease in global food production at the same time that increasing human population demands an increase in the same food supplies. Therefore, there is a great need for staple food crops that yield higher and more stably than they currently do, both in good growing years and when drought strikes. The Buckler lab for Maize Genetics and Diversity, along with the International Maize and Wheat Improvement Center (CIMMYT) Genetic Resources and Enhancement Unit is currently working to identify drought tolerance genes and alleles in maize using association mapping to build upon previous drought studies using other tools (physiology, expression profiling, comparative genetics, and QTL linkage studies).

Maize in the developing world is almost exclusively grown under rainfed conditions with minimal input and management by the small scale farmers that grow it. Although drought can strike at any time, the plants are most prone to damage due to limited water during flowering time. There are many traits that can lead to a more robust plant under moisture limited conditions, but synchrony between male and female flowering time is particularly important. This synchrony is called the Anthesis Silking Interval (ASI). Male and female flowers are physically separated, and silks are particularly prone to desiccation. Plants may delay silk production after pollen shed, but the longer the delay, the fewer grains develop. In addition to ASI, the regulation of carbohydrates are of interest to researchers working on drought tolerance, because the diminished supply of carbohydrates to the developing floral and seed organs that occurs under drought reduces seed set. Again, stress at early kernel development has a much greater negative effect on final yield than stress at a later stage (kernel filling). Abundant evidence shows that absissic acid (ABA) is involved in turning on many stress-responsive genes (Figure 1), and that it plays a key role in cell growth regulation, especially during flowering. Therefore, genes involved in ASI, genes in the carbohydrate production pathway, and genes in the ABA production pathway or genes affected by ABA itself, can be very important for the development of a drought resistant maize plant (Figure 2).

Figure 1 Role of ABA in drought response. Abscisic Acid (ABA) is a regulatory molecule involved in drought stress tolerance. The main function of ABA is to regulate plant water balance through guard cells, and regulate osmotic stress tolerance via cellular dehydration tolerance genes. In addition to drought stress, ABA is also induced by salt, and to a lesser degree, cold stress. ABA-inducible genes have the ABA-responsive element (ABRE) (C/T)ACGTGGC in their promoters.  Figure adopted from University of Toronto.

Figure 2 When drought affects a maize plant, resistant varieties will sense the stress and react by turning on key genes in pathways that will allow kernals to develop despite the limitations in water. Sensing of the limited water conditions may cause ABA to turn on genes (for example, invertase or IVR 1) that will, in turn, regulate the pathway that produces carbohydrates (sugars), particularly in the kernel. Drought resistant maize varieties will also keep a short interval between pollen shed and the emergence of silks to take the pollen to the developing kernals, while drought susceptible varieties will be unable to produce the silks in time to take advantage of pollen shed. These genes, working together in the maize plant, cause a good yield despite limited soil moisture. The ability to identify these genes will increase the efficiency of maize improvement for drought prone environments.

Once target pathways have been identified, choosing the genes within these pathways that may have the greatest effect on the desired phenotype (i.e., drought tolerant plants) becomes an important task. The sequences of these genes in the sample of individual plants under study, or just key polymorphisms within the genes, are used in studies associating the DNA sequence differences with differences in the level of drought tolerance in the study sample. Candidate genes in this study have been chosen based on: 1) genes encoding components of metabolic or signaling pathways that have established roles in carbohydrate (Figure 3) or ABA sub-systems (Figure 4), possibly in other plant species; 2) genes whose expression in previous transcript profiling experiments responded differently to drought in contrasting genotypes (individuals); and 3) genes whose map position are known and who co-localize with QTLs previously detected in maize under drought.

Figure 3 Sucrose metabolism in maize. Sucrose transported into the maize kernel is converted to glucose and fructose by the major isoform of sucrose synthase. Glucose-6-phosphate isomerase and phosphogulcomutase glucose into glucose-1-phosphate, the substrate for starch synthases. Starch synthases then sequentially add glucose-1-phosphate molecules onto the ends of a growing starch chain. Starch synthase produces amylose. The starch branching enzyme hydrolyzes linkages in amalose and reattaches them to branch points found in amylopectin.

Figure 4 The ABA biosynthetic pathway in plants. (a) Carotenoid precursor synthesis in the early steps of ABA biosynthesis. ABA is synthesized from C40 carotenoids (phytoene, _-carotene, lycopene, and _-carotene). Carotenoids are synthesized from a C5 compound, IPP. In plastids, IPP is synthesized via DXP from glyceraldehyde-3-phosphate and pyruvate. (b) Formation of epoxycarotenoid and its cleavage in plastid. (c) Reactions in the cytosol for the formation of ABA (three possible pathways are proposed).

Although the ultimate phenotype we wish to improve is grain yield of the maize plant under drought conditions, a series of correlated phenotypes are being measured to test associations against. All contribute to drought tolerance, and they include:

• Precocity
• Plant size
• Chlorophyll content
• Root conductivity
• Glucose, Sucrose, Dehydrin, ABA and ABA-glucose ester measured on leaves, ear tips, and silks harvested at different developmental stages
• Grain yield and yield components

This work was generously funded by the Generation Challenge Program.