Specialized metabolites have important roles as infochemicals in inter- and intraspecific interactions of insects. A particularly abundant class of specialized metabolites are terpenes, which are released by many members of taxonomically diverse insect lineages as pheromone and defense compounds. Despite the broad occurrence of terpenes in insects, knowledge of their biosynthesis remains limited compared to that in other forms of life. Terpenes are biosynthetically produced by the action of terpene synthase (TPS) enzymes. While insects lack TPS enzymes found in plants and microbes, there is growing evidence that insect TPS proteins have evolved independently from isoprenyl diphosphate synthase (IDS) enzymes in core terpene metabolism. To gain deeper insight into the transition from IDS to TPS function, I have explored the genomic and functional evolution of TPS enzymes in representatives of major insect lineages. First, I investigated evolutionary and functional relationships of TPS enzymes with roles in pheromone biosynthesis in pentatomids (stink bugs) including the invasive and economically critical pests Nezara viridula (Southern green stink bug) and Halyomorpha halys (brown marmorated stink bug). I also performed a comprehensive phylogenetic analysis of TPS genes in species across the broader order of piercing-sucking insects (Hemiptera), which provided evidence for an ancient emergence of TPS function in this group of insects. To gain a better understanding of core structural determinants of insect TPS evolution, we next defined distinct IDS catalytic motifs that are consistently substituted in enzymes with TPS function. These sequence characteristics were used to make predictions of TPS functionality in a large dataset of insect proteins. I determined the evolutionary dynamics of predicted and known TPS and IDS enzymes through extensive phylogenetic analysis to make top-level inferences about the distribution and evolution of TPS function in insects. Using this knowledge, I further explored functional transitions and subfunctionalization of TPS genes in the large order of beetles (Coleoptera), and more specifically, in species of the lady beetle family (Cocinellidae) including the globally invasive pest, Harmonia axyridis. Comparative genome analyses and IDS/TPS gene functional characterizations revealed gene duplication patterns and enzyme transitions that suggest TPS function evolved in part through processes of subfunctionalization and bifunctional enzymatic states. Additionally, this study provided the first experimental evidence for the mitochondrial localization of terpene metabolism in insects. Lastly, I identified putative TPS enzymes in the American cockroach, Periplaneta americana, and conducted an investigation into their catalytic activity. I found first evidence for TPS enzymatic activity in Blattodea as the most anciently diverging order of terpene-emitting insects and made inferences on the relationship of these enzymes to characterized IDS and TPS proteins in other insects. Our findings in the American cockroach point to the potential independent evolution of TPS function in blattodean cockroaches and termites in types of IDS ancestors. This work significantly advances our understanding of the evolution, functional diversity, and biochemical properties of TPS enzymes in insects, highlighting their recurring pattern of parallel evolution from IDS ancestors and its significance as a model for the emergence of novel specialized functions in core metabolic enzymes. / Doctor of Philosophy / Insects use many types of chemicals for purposes of communication and defense. Terpenes represent a common and diverse class of natural chemicals, which are used by insects to send pheromone signals and to protect themselves from predators. Terpenes also occur in other kingdoms of life. For example, in plants, they are especially widespread, forming a large portion of their essential oil and floral scent compounds. In contrast to plants and other organisms, not much is known about how insects produce terpenes. Terpenes are made by proteins called terpene synthase (TPS) enzymes. TPS enzymes have been traditionally associated with plants and microbes but have not been found in any insect species. Instead, there is growing evidence that insects have developed their own versions of these enzymes, known as isoprenyl diphosphate synthase (IDS)-type TPS enzymes, from proteins with essential functions in metabolism.
To learn more about these unique insect enzymes, we explored their evolution and activity in species of several different groups of insects. First, we investigated TPS enzymes that are required for the biosynthesis of pheromones in stink bugs including two agriculturally important pests, the Southern green stink bug and the brown marmorated stink bug. This research showed that the ability to make terpenes might be quite ancient in this group of insects compared to TPS enzymes in other insects. Next, we examined the protein structure of insect TPS enzymes and determined features that are characteristic for these types of enzymes. This information was used to predict the occurrence of TPS proteins and their evolution across many different groups of insects. In particular, I found evidence for the emergence of TPS enzymes in lady beetles, with a focus on the invasive Asian lady beetle, which emits a terpene pheromone for aggregation. My research suggested that lady beetle TPS enzymes evolved through a process called subfunctionalization, where genes duplicate and progressively split their ancestral functions with new features to evolve novel functions. This study also provided the first evidence that insects might produce terpenes in their mitochondria, a part of the cell known for energy production. Finally, I discovered potential TPS enzymes in the American cockroach. My investigation showed that cockroaches and termites, both part of the oldest-diverging group of terpene-releasing insects, may have independently developed their own TPS enzymes from different ancestor proteins. Overall, this research helps us understand how insects produce chemical compounds important to their biology and ecology and how these abilities have evolved over time. This knowledge can be useful in agriculture, pest control, and for our understanding of insect biology.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/121107 |
| Date | 10 September 2024 |
| Creators | Rebholz, Zarley Alexander |
| Contributors | Biological Sciences, Tholl, Dorothea Berta Christine, McGlothlin, Joel W., Marek, Paul, Jelesko, John G. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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