Genome-wide association mapping is highly sensitive to environmental changes, but network analysis allows rapid causal gene identification.
Genome-wide association (GWA) is gaining popularity as a means to study the architecture of complex quantitative traits, partially due to the improvement of high-throughput low-cost genotyping and phenotyping technologies. Glucosinolate (GSL) secondary metabolites within Arabidopsis spp. can serve as a model system to understand the genomic architecture of adaptive quantitative traits. GSL are key anti-herbivory defenses that impart adaptive advantages within field trials. While little is known about how variation in the external or internal environment of an organism may influence the efficiency of GWA, GSL variation is known to be highly dependent upon the external stresses and developmental processes of the plant lending it to be an excellent model for studying conditional GWA.
To understand how development and environment can influence GWA, we conducted a study using 96 Arabidopsis thaliana accessions, >40 GSL phenotypes across three conditions (one developmental comparison and one environmental comparison) and ∼230,000 SNPs. Developmental stage had dramatic effects on the outcome of GWA, with each stage identifying different loci associated with GSL traits. Further, while the molecular bases of numerous quantitative trait loci (QTL) controlling GSL traits have been identified, there is currently no estimate of how many additional genes may control natural variation in these traits. We developed a novel co-expression network approach to prioritize the thousands of GWA candidates and successfully validated a large number of these genes as influencing GSL accumulation within A. thaliana using single gene isogenic lines.
Together, these results suggest that complex traits imparting environmentally contingent adaptive advantages are likely influenced by up to thousands of loci that are sensitive to fluctuations in the environment or developmental state of the organism. Additionally, while GWA is highly conditional upon genetics, the use of additional genomic information can rapidly identify causal loci en masse.
Understanding how genetic variation can control phenotypic variation is a fundamental goal of modern biology. A major push has been made using genome-wide association mapping in all organisms to attempt and rapidly identify the genes contributing to phenotypes such as disease and nutritional disorders. But a number of fundamental questions have not been answered about the use of genome-wide association: for example, how does the internal or external environment influence the genes found? Furthermore, the simple question of how many genes may influence a trait is unknown. Finally, a number of studies have identified significant false-positive and -negative issues within genome-wide association studies that are not solvable by direct statistical approaches. We have used genome-wide association mapping in the plant Arabidopsis thaliana to begin exploring these questions. We show that both external and internal environments significantly alter the identified genes, such that using different tissues can lead to the identification of nearly completely different gene sets. Given the large number of potential false-positives, we developed an orthogonal approach to filtering the possible genes, by identifying co-functioning networks using the nominal candidate gene list derived from genome-wide association studies. This allowed us to rapidly identify and validate a large number of novel and unexpected genes that affect Arabidopsis thaliana defense metabolism within phenotypic ranges that have been shown to be selectable within the field. These genes and the associated networks suggest that Arabidopsis thaliana defense metabolism is more readily similar to the infinite gene hypothesis, according to which there is a vast number of causative genes controlling natural variation in this phenotype. It remains to be seen how frequently this is true for other organisms and other phenotypes.