Browsing by Author "Argueso, Cristiana, advisor"

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Characterization and insights into the molecular mechanism of cytokinin-induced priming of plant defenses
(Colorado State University. Libraries, 2023) McIntyre, Kathryn, author; Argueso, Cristiana, advisor; Bush, Daniel, committee member; Leach, Jan, committee member; Stewart, Jane, committee member; Reddy, Anireddy, committee member
Plants have developed several mechanisms to cope with pathogenic challenges. One of these mechanisms, known as defense priming can be effective at reducing susceptibility to pathogens. Compared to unprimed plants, the immune response from primed plants, upon pathogen attack, is much stronger. This mechanism of induced disease resistance can be initiated by biological and chemical agents. The major benefit of priming is the induction of a high level of protection with considerably low fitness costs making it an attractive disease management strategy to preserve agricultural output. Recent research has demonstrated that the plant hormone cytokinin (CK) has a priming effect against biotrophic pathogens, a phenomenon referred to here as cytokinin-induced priming (CIP). This dissertation aims to gain further understanding of CIP against the hemibiotrophic bacterial pathogens Pseudomonas syringae pv. tomato (Pst) and Pseudomonas syringae pv. maculicola (Psm) in Arabidopsis thaliana (Arabidopsis) and Brassica napus, respectively as well as the necrotrophic fungal pathogen Botrytis cinerea in Arabidopsis. Chapter 2 focuses on characterizing CIP as a true priming agent by investigating the timeframe in which CIP is most effective at reducing susceptibility to Pst and Psm in both Arabidopsis and its closely related relative, B. napus and the impacts on plant growth due to CIP in these pathosystems. Moreover, we discovered that other known priming agents depend on endogenous CK signaling suggesting CK-mediated processes are involved in the priming of defense responses. The role of CK in primed defenses against B. cinerea is explored in chapter 3 where CIP is demonstrated to reduce necrotic lesion size caused by B. cinerea in a manner dependent on the JA-mediated defenses and partially on SA-mediated defenses. Transcriptome analysis revealed that during the priming stage, CK prepares the plants for pathogenic challenge through the accumulation of cellular components needed for translation and metabolites utilized for energy production and defense. Following B. cinerea inoculation, CIP suppresses defense while increasing photosynthetic-related processes. In the final chapter, molecular mechanisms are explored during CIP against Pst. Through transcriptome changes, priming by CK potentiates gene expression associated with systemic induction of defense, also known as systemic acquired resistance (SAR), following Pst challenge. Using this information, it is demonstrated that CK treatment can also induce SAR and that the known SAR inducer, L-pipecolic acid, is dependent on endogenous CK signaling. Due to the previously identified relationship between CK and source-sink relationships, amino acid transport was demonstrated to have a role in both CIP and CK-induced SAR. New agricultural practices that mitigate crop loss due to plant diseases are beneficial in terms of sustainability and economic costs. The use of CK as a priming agent offers an avenue for a new disease management strategy in that CIP protects plants against a broad range of pathogens with minimal effects on plant growth. The molecular mechanisms underlying CIP discovered here offers new insights into the relationship between plant metabolism and defense, where its exploitation could be used to create disease protection strategies.
Open Access
Characterizing the role of plant hormones during plant development and plant immunity in Arabidopsis thaliana and Solanum lycopersicum cv. Micro-Tom
(Colorado State University. Libraries, 2022) Berry, Hannah Marie, author; Argueso, Cristiana, advisor; Bush, Daniel, committee member; Bedinger, Patricia, committee member; Stasevich, Timothy, committee member
Plant hormones are major regulators of plant growth, development, and responses to biotic and abiotic stressors. Constitutive activation of immunity is commonly associated with stunted plant growth. This phenomenon, called the growth defense tradeoff, was previously thought to result from limitations in metabolic processes, where resources were redirected from plant growth toward energetically costly defense responses. However, recent studies have shown that metabolic limitations are not solely responsible for the growth defense tradeoff, and that growth and defense can be uncoupled, resulting in plants with increased immunity without compromising plant yield. While the effects on plant growth have been widely characterized in the context of constitutive immunity, developmental impacts such as changes to plant architecture, reproductive development, and leaf morphology, have been studied to a lesser extent. Cytokinin is one of nine major plant hormone families and its role in plant growth, meristematic maintenance, cell division, and senescence are widely known. In conjunction with salicylic acid (SA), a primarily defense related hormone, cytokinin acts to promote SA-dependent defense responses, thus demonstrating a role for cytokinin in plant immunity. Perception of cytokinin initiates a two-component signaling phosphorelay leading to the activation of downstream transcription factors to induce the transcription of cytokinin-responsive genes. One group of these transcription factors is the CYTOKININ RESPONSE FACTORS (CRFs), which is found in all land plant species. In Chapter 2, I show that CRFs are negative regulators of plant growth and positively regulate plant defense responses. To further elucidate the role of CRFs in growth and immunity, I quantified growth and development in crosses between constitutive immunity mutants with elevated SA and CRF overexpressing (hereafter CRFox) lines. Here, these data show that CRFox enhances the growth restriction phenotypes previously characterized in the constitutive immunity mutants. I propose a model with CRF5 at the intersection of CK and SA crosstalk, acting as a regulator of the growth defense tradeoff. How constitutive immunity alters plant development as well as plant growth is not well understood. In Chapter 3, I characterize changes in plant architecture in constitutive immunity mutants SUPPRESSOR OF NPR1-1 (NONEXPRESSER OF PATHOGENESIS RELATED GENES 1), CONSTITUTIVE 1 (snc1) and CONSTITUTIVE EXPRESSION OF PR GENES 5 (cpr5). Both snc1 and cpr5 have elevated levels of endogenous SA and elevated disease resistance. While the reduced growth phenotypes (measured as biomass) of these mutants have been characterized, phyllotaxy has not previously been quantified. Phyllotaxy describes the consistent arrangement or pattern of consecutive organs around a central point. Arabidopsis has a spiral phyllotactic pattern where each consecutive organ is separated by approximately 137.5°. Phyllotactic analysis of shoot apical meristems (SAMs) using scanning electron microscopy and the arrangement of siliques on inflorescence stems in snc1 mutants showed a change in phyllotactic divergence angle originating from reduced shoot apical meristem size and increased plastochron ratio. The plastochron describes the amount of time between organ initiations but can be shown as a ratio when the edge of two consecutive inflorescence primordia to the SAM center is quantified using imaging software. To mimic the phenotypes of constitutive immunity, I inoculated wild type Arabidopsis Col-0 plants with high concentrations of Pseudomonas syringae pv. tomato (Pst) strains: (1) Pst hrcC-, which is lacks the type-III secretion system necessary to introduce bacterial effectors into the plant cell but initiates plant basal immune responses, thus inducing pattern triggered immunity (PTI); (2) Pst DC3000, which causes plant disease via introduction of bacterial effector proteins into the plant cell to change plant metabolism and dampen plant immune responses causing effector triggered susceptibility (ETS); and (3) Pst ArvRpm1 where the bacterial effector protein Rpm1 is recognized by the plant, initiating a high level of defense responses called effector triggered immunity (ETI). Only multiple, concentrated inoculations of Pst DC3000 were able to induce changes to silique phyllotactic patterns. Notably, the SAM was unaltered after Pst DC3000 inoculations, demonstrating, that phyllotactic patterns originating at the meristem are very robust upon infection, and this change in silique patterning was determined to be a result of post-meristematic stem torsion. I conclude that elevated SA throughout the plant life in the constitutive immunity mutants shows a role for SA in regulating meristematic maintenance and/or patterning, but elevated SA after pathogen attack is not sufficient to overcome the tight regulation of the meristem. Arabidopsis has been a primary source of knowledge for elucidating hormone crosstalk during plant development and immunity. However, Arabidopsis cannot inform us about the roles of hormones during fleshy fruit development. Tomatoes are an important agricultural crop species, have a fully sequenced genome, and many genetic resources are available, making tomatoes a model crop species. In my last research chapter, I quantified plant hormones in Solanum lycopersicum cv. Micro-Tom in above- and below-ground plant tissues at four stages of plant development. I selected plant developmental stages based on easily definable traits: the seedling stage was defined as the presence of cotyledons before the emergence of true leaves; the young developmental stage was characterized by the presence of four true leaves before the transition to flowering; the adult or flowering stage was determined by the opening of the first flower; and the fruiting stage was identified by fruit set and fully expanded, breaker, and ripe fruit development stages. While the data collected in this chapter is primarily descriptive, we showed that a single extraction protocol could be used to extract and quantify 18 plant hormones representing 5 of the 9 major hormone families in multiple tissue types including roots, leaves, and fruits. Plant hormone data were integrated into botanical illustrations to create the Plant Hormone Atlas, which was presented at the Art Lab Fort Collins, in Fort Collins, CO.
Open Access
Cytokinin-mediated processes promote heat-induced disease susceptibility of plants to bacterial pathogens
(Colorado State University. Libraries, 2021) Shigenaga, Alexandra Marie, author; Argueso, Cristiana, advisor; Bush, Dan, committee member; Leach, Jan, committee member; Heuberger, Adam, committee member; Nishimura, Marc, committee member
As global human populations continue to grow and temperatures are expected to rise, the pressure to increase food productivity and develop more stress-resistant crop varieties intensifies. Increased temperatures, a consequence anticipated as a result of global climate change, is expected to have an overall negative impact on crop productivity and agricultural systems. When exposed to non-optimal, high temperature conditions plant defense responses to pathogen attack are attenuated, leading to a process referred to here as heat-induced disease susceptibility. The plant growth hormone cytokinin is known to regulate responses to both biotic and abiotic pressures, making it an ideal target to study heat-induced disease susceptibility. The overarching goal of this dissertation was to understand the role of cytokinin in heat-induced disease susceptibility, to identify novel strategies to combat this process and design new ways to teach future generations about the impact of climate change on agricultural systems and science policy. First, I identified that a plant lacking a functional cytokinin signaling pathway, ahk2,3 mutated on the cytokinin signaling receptors AHK2 and AHK3, was less susceptible at elevated temperatures to the bacterial pathogen, Pseudomonas syringae pv. tomato DC3000 (Pst DC3000). My results show that ahk2,3 plants are less susceptible under high temperature conditions with Pst DC3000 populations proliferating at a lower rate compared to wild-type plants overtime, suggesting that heat-induced susceptibility is partially dependent on cytokininiii signaling. Our results show that differences in susceptibility under elevated temperatures of ahk2,3 and wild-type plants is not attributed to an increase in defense responses, but rather by a possible change in the availability of nutrients for Pst DC3000. Together the data reveals that under high temperature conditions cytokinin promotes late-physiological processes, centered around primary metabolism, that are contributing to increased pathogen proliferation. These results led to the identification of cytokinin-regulated genes that could be utilized for breeding efforts to obtain loss-of-heat induced disease susceptibility that could be translated to crop species. Second, I identified that another member of the Brassicaceae family, Brassica napus, also exhibited heat-induced disease susceptibility to the bacterial pathogen, P. syringae pv. maculicola (Psm ES4326). Gene expression analysis confirms that similar to Arabidopsis, B. napus plants increase cytokinin signaling in response to high temperature stress. To further address if cytokinin was important for heat-induced disease susceptibility of B. napus, I utilized a chemical approach. B. napus plants were sprayed with the cytokinin-signaling antagonist, PI-55, prior to inoculation and results show that a single application of PI-55 led to a loss of susceptibility under heat to Psm ES4326. Additionally, this application of PI-55 did not lead to any adverse vegetative growth parameters, suggesting a potential novel chemical approach to combat heat-induced disease susceptibility in Brassicaceae crops. Lastly, I constructed a new approach to teach future generations about the impact of climate change on plant diseases in agricultural systems. "Plant Diseases and Climate Change" is an active learning activity designed to give college students experience in synthesizing information and developing a solution, in the context of plant pathology. This exercise uses the issue of heat-induced susceptibility of rice in the Philippines to improve student understanding of the interactions between abiotic and biotic factors affecting global food security. By using an international agricultural pathosystem, I aim to inform students how environmental pressures can impact economically important plant systems, the role scientists and experts play in policy making to preserve food security, and the importance of agriculture on a global scale.