Master of Science / Biochemistry and Molecular Biophysics Interdepartmental Program / Timothy P. Durrett / Many Euonymus species express an acetyltransferase enzyme in their seeds which catalyzes the transfer of an acetyl group from acetyl-CoA to the sn-3 position of diacylglycerol (DAG) producing unusual acetyl-1,2-diacyl-sn-glycerols (acetyl-TAG). The presence of the sn-3 acetate group gives acetyl-TAG with unique physical properties over regular triacylglycerol (TAG) found in vegetable oils. The useful characteristics of acetyl-TAG oil offer advantages for its use as emulsifiers, lubricants, and 'drop-in' biofuels. One enzyme, Euonymus alatus diacylglycerol acetyltransferase (EaDAcT), responsible for acetyl-TAG synthesis in nature was previously isolated from the seeds of Euonymus alatus (burning bush) and expressed in the oilseed crop Camelina sativa. Expression of EaDAcT successfully led to production of high levels of acetyl-TAG in camelina seeds. To further increase acetyl-TAG accumulation in transgenic camelina seeds, multiple strategies were examined in this study. Expression of a new acetyltransferase enzyme (EfDAcT) isolated from the seeds of Euonymus fortunei, which was previously shown to possess higher in vitro activity and in vivo acetyl-TAG levels compared to EaDAcT, increased acetyl-TAG accumulation by 20 mol%. Suppression of the endogenous competing enzyme DGAT1 further enhanced acetyl-TAG accumulation to 90 mol% in selected transgenic line. Studying the regulation of EfDAcT transcript, protein, and acetyl-TAG levels during seed development further provided new insights on the factors limiting acetyl-TAG accumulation.

Elevated atmospheric CO2 has been implicated as a driver of increased liana abundance worldwide. Known as disturbance creators and beneficiaries, lianas possess the potential to significantly influence forest ecosystems. I investigated the early-invasion dynamics of Euonymus fortunei (wintercreeper), an evergreen liana that is invading forests in eastern North America, disrupting native plant communities and ecosystem functions. Wintercreeper is widely cultivated as an ornamental groundcover, frequently invading natural areas via asexual stem growth. Invasion of remote natural areas is dependent upon seed transport and may occur less frequently. I examined the mechanisms of seed dormancy by conducting a ‘move-along’ experiment using fresh and after-ripened seeds. Additionally, I sought to characterize the nature of seed dispersal by birds by deploying seed traps within an invaded forest in central Kentucky. Wintercreeper seeds displayed conditional nondeep physiological dormancy. Although germination occurs at high velocity following cold stratification, a cold period was only facultative to break dormancy. While fresh seeds had greater germination rates (98.6%) compared to after-ripened seeds (85.7%), after-ripened seeds experienced earlier germination (41 days across all treatments). My findings also indicate that bird-mediated seed dispersal occurs throughout the winter from seeds that after-ripen on maternal liana phanerophytes. Overall, these data suggest wintercreeper seeds are capable of dispersing and recruiting in areas with mild to non-existent winters. The invasion of natural areas by exotic invaders is regulated by biotic and abiotic processes, which influence the invader’s success or failure. I studied the vegetation and soil effects of paired invaded (INV), uninvaded (NAT), and ‘restored’ (RES) sites on the germination and survival of wintercreeper seeds and seedlings. The effect of aril (with vs. without) was also tested, both in the field and in vitro. In the field, total germination and first year survival were 55.6% and 24.2%, respectively, across treatments. Total germination was unaffected by treatments, yet vegetation (P = 0.0016) and aril (P = 0.001) treatments significantly influenced germination rates over time, including delayed germination of seeds with arils. The proportion of germinated seedlings that survived was significantly different based on vegetation (P = 0.054) and aril (P = 0.071) treatments after the first winter of growth, but not prior to the first winter. The proportion of seedling survival was significantly lower among seeds dispersed with an aril and seeds sown within INV treatments; there were no interactive effects. Finally, I examined how seedling density and growth habit (horizontal vs. vertical) influenced plant survival, growth, and allometry. I also tested extractable soil C, N, P, K, Ca, and Mg prior to planting and after 17 months of soil conditioning. I found evidence that increased planting density negatively influenced growth among individual plants (lower survival, basal diameter, shoot mass, root mass). At the plot level, high-density plantings yielded greater stem length, and shoot, root, and combined biomass, indicating positive frequency dependence for this species. Soil analyses indicated C, N, P, Ca, and Mg significantly increased (P < 0.05) over the course of the experiment. I conclude that in wintercreeper, prominent asexual propagation leads to aggregate populations whose total contributions to above- and belowground biomass are positively correlated with density, even though individual plant fitness is not. Altogether, this work provides insight into how wintercreeper invasions occur at broad and fine scales. This information will provide a foundation for future wintercreeper studies and aid land managers in their prevention and control strategies.

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