Investigating the spatial and temporal scales of adaptation has proven difficult because the molecular basis of ecologically important traits is often complex. This project examines in white clover the process of adaptation using five independent geographical gradients in a well-characterized trait called cyanogenesis (cyanide release after tissue damage). Specifically, this work compares genetic variation throughout the clover genome to variation in the two genes underlying cyanogenesis, with the goal of understanding the degree to which natural selection, gene flow, and other processes are conserved across gradients. The origins of the adaptive variation in each gradient will also be investigated. Further, ecological experiments will be used to identify the ecological trade-offs that maintain cyanogenesis variation within populations.
Understanding the processes that create and maintain ecologically important variation is fundamental in making predictions regarding species responses to changing environments and establishing future policy. This proposal investigates these processes in a tractable natural system. This work will answer broad questions about adaptation while providing a useful case study for predicting species responses. This proposal carries a large educational outreach component for undergraduates and high school students, using clover cyanogenesis as a basic teaching tool for understanding the connection between genetics and ecology. The project will also provide training to young scientists and high school teachers in laboratory and field settings.
Clines, changes in a trait or allele frequency over an environmental gradient, are frequently used to study the process of adaptation in environmentally variable areas. If clines in a particular trait occur recurrently in different geographical areas, they can be used as independent replicates to examine the predictability of climatic adaptation. This PhD dissertation project examines clines in cyanogenesis, the ability to produce toxic hydrogen cyanide after tissue damage, in plants. In white clover, some plants are cyanogenic while others are acyanogenic. This polymorphism is often spatially distributed in clines, where the frequency of cyanogenic plants is higher in populations found in warmer climates. Previous studies suggest that populations growing in locations with milder winters have a high abundance of herbivores, and that cyanogenesis clines develop because cyanogenesis is an effective but energetically costly herbivore deterrent. This project sampled clines spanning similar environmental gradients in different areas of the world where white clover has been introduced to examine whether similar selective forces have acted on the same genes to create similar clinal patterns. Analyses indicate that the mechanisms of cline formation differ among regions. Two necessary precursors for cyanogenesis, cyanogenic glucosides and linamarase, may be either present or absent in a given plant, and the frequencies of these biochemical precursors show different patterns among regions. For instance, in an altitudinal transect along Mt. Baker (Washington state), there is a cline in the frequency of plants with cyanogenic glucosides, with more plants with cyanogenic glucosides at lower elevations. However, in the Southern Alps of New Zealand, there is only a cline in the frequency of plants producing linamarase, not cyanogenic glucosides. Our analyses indicate that these different patterns are likely attributable to functions for cyanogenesis other than herbivore defense. For example, growth chamber experiments indicate that plants with cyanogenic glucosides are able to better withstand drought stress, suggesting adaptation to dry climates. Data also indicate that the molecular genetic basis of cyanogenesis variation is shared among regions. This suggests that the mutations underlying this variation most likely predate the introduction of white clover into its non-native range. More generally, the project results indicate that adaptation can occur rapidly after introduction of a species into a new region, that mechanisms of adaptation are context-dependent, and that recurrent patterns in nature may arise through multiple interacting ecological factors. The broader impacts of this doctoral dissertation project included mentoring of high school students and undergraduates, as well as developing a classroom lab activity for high school and college Biology classes. This activity allows students to collect clover plants in backyards and parks, test them for cyanogenesis in the classroom, and compare their results to classes in other parts of the country; it thus provides a hands-on lab activity that connects genetics to ecology and climatic adaptation. During the course of this project, the PI and co-PI gave presentations on the clover classroom lab activity to >100 biology teachers at conferences, summer institutes, and visits to local high school biology classes in St. Louis. A total of three undergraduate researchers and one high school student also received training through this project; they helped design experiments, conducted lab and field work, and performed statistical data analysis. This work resulted in two presentations at conferences by undergraduates, as well as multiple scientific presentations and publications by the project PI and co-PI.