Flowering plants display diverse reproductive systems, including a variety of gender strategies and mechanisms of clonal propagation. Here, I investigate gender strategies, sex-ratio evolution, and sexual dimorphism in the North American clonal aquatic, Sagittaria latifolia (Alismataceae), which exhibits three sex phenotypes (hermaphrodites, females, males) and two modal sexual systems (monoecy, dioecy). This provides an outstanding opportunity to examine the costs and benefits of combined versus separate sexes. My research focused on the northern range limit in eastern N. America, and on disjunct populations in western N. America. I developed microsatellite (SSR) markers to investigate population genetic structure at several spatial scales, including the clonal structure of local populations to continental patterns. These analyses provided insights on the roles of historical, ecological and reproductive factors in the evolution and maintenance of sexual system diversity. Phenotypic sex ratios varied near continuously from monoecy through subdioecy (three sex phenotypes) to dioecy. A comparison of phenotypic and genotypic sex ratios in dioecious populations demonstrated close correspondence. The northern range limit was characterized by a decline in female frequency and an increased incidence of subdioecy. I evaluated two hypotheses to explain this pattern: (1) increased sex inconstancy in dioecious populations; (2) hybridization between monoecious and dioecious populations. I found support for both hypotheses, although hybridization appears to be the more common pathway to subdioecy. I parameterized a model predicting female frequency and hermaphrodite sex allocation; observed and predicted values were correlated suggesting that subdioecious populations are closer to equilibrium than expected for a clonal perennial. A comparison of eastern and western populations indicated genetic differentiation between monoecy and dioecy in the east, but in the west, due to habitat isolation, geography plays a more important role in genetic differentiation. Evidence from cpDNA haplotype variation indicated that the western range was established following long-distance colonization from the east involving a genetic bottleneck. The discovery of gynodioecious populations of S. latifolia in the west, and the absence of ecological and genetic differentiation between monoecious and dioecious populations, raise the possibility that dioecy may have evolved autochthonously in the west, and more recently than in the eastern range.

Seed formation consists of two major stages: embryo pattern formation and maturation. During seed maturation, the embryo accumulates storage material, acquires desiccation tolerance, and enters a stage of dormancy. Genetic analyses have identified several master regulators that orchestrate late embryogenesis, including the B3-domain transcription factor FUSCA3 (FUS3). In Arabidopsis, FUS3 has been shown to be a central regulator of hormonal pathways; it positively regulates late embryogenesis by increasing abscisic acid (ABA) level while repressing gibberellin (GA) synthesis. In turn, FUS3 protein level is positively and negatively regulated by ABA and GA, respectively. However, the mechanism of how this regulation occurs has not been well characterized. In this study, FUS3 has been shown to be an unstable protein rapidly degraded by the proteasome through a PEST instablility motif. To further characterize the mechanisms involved in FUS3 homeostasis, FUS3-interacting proteins were identified. The SnRK1 kinase AKIN10 was shown to interact with and phosphorylate FUS3 at its N-terminus. Furthermore, overexpression of AKIN10 delays FUS3 degradation, suggesting AKIN10 positively regulates FUS3 protein accumulation. Overexpression of AKIN10 delays developmental phase transitions, and causes defects in lateral organ development. These defects were partially rescued by the loss-of-function fus3-3 mutation, suggesting FUS3 and AKIN10 genetically interact to regulate these developmental processes. SnRK1/AMPK/Snf1 kinases are regulators of energetic stress responses. Overexpression studies suggest both FUS3 and AKIN10 positively regulate ABA signaling, but differ in sugar responses during germination; AKIN10 mediates glucose sensitivity, while FUS3 regulates osmotic stress responses. Overexpression of AKIN10 and FUS3 results in glucose and osmotic stress hypersensitivities, respectively, both of which are partially dependent on de novo ABA synthesis. Thus, FUS3 and AKIN10 act in overlapping pathways and combine different environmental signals to generate a common ABA-dependent response. In summary, novel mechanisms that regulate FUS3 homeostasis and function were identified. A model explaining the interaction between FUS3 and AKIN10 during embryonic and vegetative development, and the function of these two central developmental regulators in hormonal and stress signaling pathways is discussed.

We present the first characterization of K+ optimization of N uptake and metabolism in an NH4+-tolerant species, tropical lowland rice (cv. IR-72). 13N radiotracing showed that increased K+ supply reduces futile NH4+ cycling at the plasma membrane, diminishing the excessive rates of both unidirectional influx and efflux. Pharmacological testing showed that low-affinity NH4+ influx may be mediated by both K+ and non-selective cation channels. Suppression of NH4+ influx by K+ occurred within minutes of increasing K+ supply. Increased K+ reduced free [NH4+] in roots and shoots by 50-75%. Plant biomass was maximized on 10 mM NH4+ and 5 mM K+, with growth 160% higher than 10 mM NO3--grown plants, and 220% higher than plants grown at 10 mM NH4+ and 0.1 mM K+. Unlike in NH4+-sensitive barley, growth optimization was not attributed to a reduced energy cost of futile NH4+ cycling at the plasma membrane. Activities of the key enzymes glutamine synthetase (GS) and phosphoenolpyruvate carboxylase (PEPC) were strongly stimulated by elevated K+, mirroring plant growth and protein content. Improved plant performance through optimization of K+ and NH4+ is likely to be of substantial agronomic significance in the world’s foremost crop species.

To improve fungal identification using a single DNA sequence, I introduce the Barcode Similarity Group (BSG) defined as a cluster of sequences that share greater than or equal to a threshold amount of genetic similarity with each other. As a test case, I created 393 BSGs from 2463 Cortinarius ITS sequences using a 94% similarity cut-off value in DOTUR. Some BSGs may contain multiple species. The BSG database was used to label environmental sequences, find misidentified or mislabeled sequences, and find potential cryptic species and novel species. Expert taxonomists will be needed to perform detailed morphological and phylogenetic studies to identify the individual species within each BSG. The main advantage of using BSGs is that it clusters together sequences using total genetic relatedness and does not rely on any taxonomy for identification. A website was created where the RDP Classifier is used to classify a query sequence into a BSG.

Flowering plants display diverse reproductive systems, including a variety of gender strategies and mechanisms of clonal propagation. Here, I investigate gender strategies, sex-ratio evolution, and sexual dimorphism in the North American clonal aquatic, Sagittaria latifolia (Alismataceae), which exhibits three sex phenotypes (hermaphrodites, females, males) and two modal sexual systems (monoecy, dioecy). This provides an outstanding opportunity to examine the costs and benefits of combined versus separate sexes. My research focused on the northern range limit in eastern N. America, and on disjunct populations in western N. America. I developed microsatellite (SSR) markers to investigate population genetic structure at several spatial scales, including the clonal structure of local populations to continental patterns. These analyses provided insights on the roles of historical, ecological and reproductive factors in the evolution and maintenance of sexual system diversity. Phenotypic sex ratios varied near continuously from monoecy through subdioecy (three sex phenotypes) to dioecy. A comparison of phenotypic and genotypic sex ratios in dioecious populations demonstrated close correspondence. The northern range limit was characterized by a decline in female frequency and an increased incidence of subdioecy. I evaluated two hypotheses to explain this pattern: (1) increased sex inconstancy in dioecious populations; (2) hybridization between monoecious and dioecious populations. I found support for both hypotheses, although hybridization appears to be the more common pathway to subdioecy. I parameterized a model predicting female frequency and hermaphrodite sex allocation; observed and predicted values were correlated suggesting that subdioecious populations are closer to equilibrium than expected for a clonal perennial. A comparison of eastern and western populations indicated genetic differentiation between monoecy and dioecy in the east, but in the west, due to habitat isolation, geography plays a more important role in genetic differentiation. Evidence from cpDNA haplotype variation indicated that the western range was established following long-distance colonization from the east involving a genetic bottleneck. The discovery of gynodioecious populations of S. latifolia in the west, and the absence of ecological and genetic differentiation between monoecious and dioecious populations, raise the possibility that dioecy may have evolved autochthonously in the west, and more recently than in the eastern range.

Seed formation consists of two major stages: embryo pattern formation and maturation. During seed maturation, the embryo accumulates storage material, acquires desiccation tolerance, and enters a stage of dormancy. Genetic analyses have identified several master regulators that orchestrate late embryogenesis, including the B3-domain transcription factor FUSCA3 (FUS3). In Arabidopsis, FUS3 has been shown to be a central regulator of hormonal pathways; it positively regulates late embryogenesis by increasing abscisic acid (ABA) level while repressing gibberellin (GA) synthesis. In turn, FUS3 protein level is positively and negatively regulated by ABA and GA, respectively. However, the mechanism of how this regulation occurs has not been well characterized. In this study, FUS3 has been shown to be an unstable protein rapidly degraded by the proteasome through a PEST instablility motif. To further characterize the mechanisms involved in FUS3 homeostasis, FUS3-interacting proteins were identified. The SnRK1 kinase AKIN10 was shown to interact with and phosphorylate FUS3 at its N-terminus. Furthermore, overexpression of AKIN10 delays FUS3 degradation, suggesting AKIN10 positively regulates FUS3 protein accumulation. Overexpression of AKIN10 delays developmental phase transitions, and causes defects in lateral organ development. These defects were partially rescued by the loss-of-function fus3-3 mutation, suggesting FUS3 and AKIN10 genetically interact to regulate these developmental processes. SnRK1/AMPK/Snf1 kinases are regulators of energetic stress responses. Overexpression studies suggest both FUS3 and AKIN10 positively regulate ABA signaling, but differ in sugar responses during germination; AKIN10 mediates glucose sensitivity, while FUS3 regulates osmotic stress responses. Overexpression of AKIN10 and FUS3 results in glucose and osmotic stress hypersensitivities, respectively, both of which are partially dependent on de novo ABA synthesis. Thus, FUS3 and AKIN10 act in overlapping pathways and combine different environmental signals to generate a common ABA-dependent response. In summary, novel mechanisms that regulate FUS3 homeostasis and function were identified. A model explaining the interaction between FUS3 and AKIN10 during embryonic and vegetative development, and the function of these two central developmental regulators in hormonal and stress signaling pathways is discussed.

We present the first characterization of K+ optimization of N uptake and metabolism in an NH4+-tolerant species, tropical lowland rice (cv. IR-72). 13N radiotracing showed that increased K+ supply reduces futile NH4+ cycling at the plasma membrane, diminishing the excessive rates of both unidirectional influx and efflux. Pharmacological testing showed that low-affinity NH4+ influx may be mediated by both K+ and non-selective cation channels. Suppression of NH4+ influx by K+ occurred within minutes of increasing K+ supply. Increased K+ reduced free [NH4+] in roots and shoots by 50-75%. Plant biomass was maximized on 10 mM NH4+ and 5 mM K+, with growth 160% higher than 10 mM NO3--grown plants, and 220% higher than plants grown at 10 mM NH4+ and 0.1 mM K+. Unlike in NH4+-sensitive barley, growth optimization was not attributed to a reduced energy cost of futile NH4+ cycling at the plasma membrane. Activities of the key enzymes glutamine synthetase (GS) and phosphoenolpyruvate carboxylase (PEPC) were strongly stimulated by elevated K+, mirroring plant growth and protein content. Improved plant performance through optimization of K+ and NH4+ is likely to be of substantial agronomic significance in the world’s foremost crop species.

To improve fungal identification using a single DNA sequence, I introduce the Barcode Similarity Group (BSG) defined as a cluster of sequences that share greater than or equal to a threshold amount of genetic similarity with each other. As a test case, I created 393 BSGs from 2463 Cortinarius ITS sequences using a 94% similarity cut-off value in DOTUR. Some BSGs may contain multiple species. The BSG database was used to label environmental sequences, find misidentified or mislabeled sequences, and find potential cryptic species and novel species. Expert taxonomists will be needed to perform detailed morphological and phylogenetic studies to identify the individual species within each BSG. The main advantage of using BSGs is that it clusters together sequences using total genetic relatedness and does not rely on any taxonomy for identification. A website was created where the RDP Classifier is used to classify a query sequence into a BSG.

No description available.

Doctor of Philosophy / Department of Horticulture, Forestry, and Recreation Resources / Dale J. Bremer / Turfgrass covers 1.9% of the nation’s surface area and is the largest irrigated crop in the USA. Developed urbanized land is projected to double by 2025, which will increase turf’s environmental impact. Studies were conducted to evaluate environmental impacts by characterizing nitrogen, carbon, and water fluxes in turfgrass ecosystems. Emissions of nitrous oxide (N[subscript]2O), a major greenhouse gas and ozone depleter were measured from bermudagrass (Cynodon dactylon L. Pers. x C. transvaalensis Burtt-Davy) (bermuda), perennial ryegrass, (Lolium perenne L.) (rye), and zoysiagrass, (Zoysia japonica Steud.) (zoysia) under regional N management. In a separate study, N2O fluxes were measured from bermuda fertilized with controlled-release N fertilizers including polymer-coated and organic-N, and quick release urea. Emissions of N2O were measured using static surface chambers and gas chromatography. Zoysia, with less N requirements, had lower emissions than bermuda. Cumulative N[subscript]2O emissions were similar among N types. To measure water and carbon fluxes, a portable non-steady state chamber was designed and tested. The chamber had minimal affects to the canopy during field measurements: leak values averaged <1.5 micromol CO[subscript]2 m[superscript]-2 s[superscript]-1; average chamber pressure was 0.09 Pa ±0.01 Pa; temperature rise inside the chamber averaged 0.74C; and the chamber had 90% photosynthetically active radiation transmittance. Using the chamber, differences were detected in net photosynthesis (Pnet), gross photosynthesis (Pg), evapotranspiration (ET), canopy stomatal conductance (gc), and water use efficiency (WUE) in well-watered tall fescue (Festuca arundinacea Schreb.), Kentucky bluegrass (Poa pratensis L.) (KBG), zoysia, and bermuda. Irrigation requirements, visual quality ratings, and genetic rooting potential of 28 KBG cultivars and 2 Texas bluegrass hybrids (P. pratensis x P. arachnifera Torr.) were quantified in greenhouse and rainout facility studies. Average water applied ranged from 23.4 to 40.0 cm among cultivars. Bedazzled, Preakness, and Bartitia required less water and had higher average quality than other cultivars. Compact America and Mid-Atlantic phenotypes exhibited greatest potential for success in integrating reduced water inputs with maintenance of acceptable visual quality. Results indicated that turfgrass management could mitigate N[subscript]2O emissions and conserve water while maintaining healthy turfgrass, and the new chamber will enhance turfgrass studies by providing rapid measurements of photosynthesis.