2016 Food & Fuel for the 21st Century Symposium

La Jolla, California
Thursday, March 10, 2016
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POSTER ABSTRACTS

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Identifying novel regulators of maize herbivore-associated volatile terpene biosynthesis through phosphoproteomic profiling

Alisa Huffaker, Philipp Weckwerth, Elly Poretsky, Andrew Sher, Zhouxin Shen, Steven Briggs
UC San Diego

Maize (Zea mays) is a model for the study of herbivore-induced production of terpene volatiles that attract parasitic wasps, aiding in plant protection. Recently a peptide signal, ZmPep3, was demonstrated to be a potent regulator of maize herbivory-associated volatiles, stimulating expression of terpene synthase genes and emission of a volatile blend similar to those observed upon caterpillar attack. ZmPep3 is as potent as insect-derived elicitors in promoting these defense responses and thus a useful tool for probing molecular mechanisms controlling herbivore-associated terpene production. To better understand molecular events regulating inducible volatile terpene biosynthesis, we have characterized changes in the maize proteome and phosphoproteome triggered by ZmPep3.  12 hours post-treatment, ZmPep3 caused accumulation of numerous proteins associated with terpene biosynthesis, including terpene synthases, farnesyl diphosphate synthase (FPS) and 1-deoxy-D-xylulose-5-phosphate synthase (DXS).  Within 10 minutes ZmPep3 treatment induced significant changes in phosphorylation of signaling proteins such as transcription factors, kinases, receptors and RNA binding proteins.  Several of these candidates are orthologs of proteins demonstrated to promote anti-herbivore responses in other plants, but many are novel elements that are being investigated for their regulatory role with regards to volatile terpene biosynthesis and defense against herbivores.

A Novel Broad Spectrum Extraction Method for the Simultaneous Isolation and Detection of Defense-related Metabolites and Phytohormones

Chan, Joshua; Christensen, Shawn; Luu, An; Ding, Yezhang; Schmelz, Eric
UC San Diego

Plant secondary metabolites constitute a wide range of biosynthetic classes with a vast array of properties that critically influence plant defenses, crop resilience and human health. Given the chemical diversity, current conventional extraction methods are largely unsatisfactory for the simultaneous detection and quantification of labile defense-related metabolites alongside trace analytes such as phytohormones. To address this need, we developed an extraction procedure utilizing a novel combination of solid-liquid and liquid-liquid extraction combined with LC-MS (QQQ) analyses to characterize broad metabolomic changes. In the present study, we use this method to explore the inducible defense responses of maize tissues challenged with a fungal pathogen (Cochliobolus heterostrophus; Southern corn leaf blight). Inoculated maize stems revealed pathogen elicited increases in 565 significant (p ≤ 0.05) features considering both positive (288) and negative (277) ionization modes, respectively. Highly induced features are comprised largely of acidic terpenoids, anthocyanidins, flavonoids, flavonoid conjugates, phenylpropanoids, phenolic amides, non-protein amino acids, benzoxazinoids, and phytohormones. The results of our newly developed method uniquely allows for the isolation and quantification of vastly disparate classes of defense related compounds that have not previously been considered together and enables plant defense metabolism to be examined on a system wide level.

New Insights On The Genetics, Biochemistry, Diversity And Function Of Maize Defenses

Ding, Yezhang; Chan, Josh; Huffaker; Alisa; Schmelz, Eric
UC San Diego

As the world’s largest annually harvested crop, maize has become a leading research model to understand the biochemical mechanisms that mediate plant resilience to both biotic and abiotic stress. A quarter of a century ago, it was discovered that herbivory strongly elicits maize sesquiterpene volatile emission which in turn can function as host location signals for predators and parasitoids. In addition to well-studied herbivore-induced maize defenses, pathogens also strongly elicit a subset of terpene synthases (ZmTPS) often in the absence of detectable increases in anticipated volatile olefins. Following fungal infection, β-macrocarpene synthases (ZmTPS6/11) and the ent-copalyl diphosphate synthase (ZmAn2) are among the most highly inducible transcripts in maize and mediate the production of locally accumulated antimicrobial phytoalexins, termed zealexins and kauralexins. Recent metabolomic, transcriptomic, QTL and mutant analyses have revealed a far greater diversity of defense-related biochemicals, genes and functions than previously appreciated. Minimally six pathogen inducible ZmTPS yield products that are predictably oxidized by cytochrome P450s and contribute to a complex suite of bioactive molecules in pathogen challenged tissues. As an example of previously hidden metabolic diversity, we detail the recent QTL mapping of α/β-selinene biosynthesis to candidate TPS and P450 gene clusters responsible for a predominant phytoalexin pathway leading to α/β-costic acids. Beyond terpenoids, maize phytoalexins span benzoxazinoids, flavonoids, phenolics, and oxylipins all of which likely generate select metabolites that additionally act as multifunctional signals regulating plant transcriptional responses. The combination of metabolomic, transcriptomic, and QTL approaches is a powerful tool to rapidly uncover novel maize defense pathways and enable the functional annotation of previously unknown genes mediating disease resistance.

Characterization of downstream Pep signaling components: importance of a Pep-responsive RNA recognition motif (RRM)-containing protein

Dressano, Keini; Huffaker, Alisa
UC San Diego

The human population growth will require an increasing of agricultural productivity. Strategies to enable such increase are necessary to improve yield, minimizing the loss caused by pests and pathogens, which is a problem in production of important crops. Plants are able to perceive molecules from foreign organisms as microbes to detect biotic attack, and in response to that stimulus, they can produce and recognize endogenous peptide elicitors related to the immune response. The Plant Elicitor Peptides (Peps) were first discovered in Arabidopsis plants and regulate defense responses, enhancing plant resistance to pathogens. The orthologous Pep peptide- and receptor-encoding genes were discovered in over 90 plant species and all major crops. In maize, the orthologous ZmPep1 mediates pathogen resistance. Interestingly, ZmPep3 regulates direct and indirect anti-herbivore defense responses, showing that Peps are involved in defense responses against herbivores as well. Although Peps have shown to be fundamental plant immunoregulatory signals, the signal transduction pathway for these peptides is still poorly understood. The identification and characterization of Pep signaling pathway components will contribute to elucidate how Peps induce resistance to pathogens and herbivores, and it will also allow the development of powerful strategies for improvement of plant immunity. To identify Pep signaling components, the profiling of rapid changes in the phosphoproteome was investigated in suspension-cultured cells of Arabidopsis and maize treated with AtPep1 and ZmPep3, respectively. One of the proteins significantly altered in phosphorylation state in response to both treatments was an RNA recognition motif (RRM)-containing protein. The main aim of this work is to characterize the molecular mechanisms by which this RNA recognition motif (RRM)-containing protein affects Pep signaling and immunity. Our results showed that mutants containing T-DNA insertion in gene encoding this protein are hypersensitive to AtPep1, indicating the potential activity of RRM as a negative regulator of AtPep1-induced responses.   

Induced foliar volatile production in response to the herbivore elicitor N-linolenoyl L-glutamine maps to a single QTL in maize

Elly Poretsky, Philipp Weckwerth, Eric Schmelz, Alisa Huffaker
UC San Diego

The fatty acid amide elicitor N-linolenoyl L-glutamine (Gln-18:3) is present in the oral secretions of Lepidopteran herbivores and is a potent inducer of vegetative volatile emissions associated with indirect defense against herbivores. Maize plants treated with Gln-18:3 emit a rich chemical blend, including numerous terpenes and green leafy volatiles (GLV). Despite the potent volatile-inducing activity of Gln-18:3, sensitivity to this elicitor varies widely among species and even among cultivars within the same species. To understand the molecular phenotype underlying plant competency to respond to Gln-18:3, better characterization of signaling components specific to the elicitor are needed. We identified two parent maize plants with differential abilities to respond to Gln-18:3, but similar responsiveness to other elicitors of volatile emissions. A recombinant inbred line (RIL) population produced from these two parents was screened for sensitivity to Gln-18:3 through measurement of induced volatile emission upon treatment. Mapping of the response patterns revealed a single quantitative trait locus (QTL) that is significantly associated with emission of both terpene and GLVs after treatment with Gln-18:3. Candidate genes at this locus have been identified and their potential role in modulating sensitivity to Gln-18:3 is being examined.

More than a model: High-yield growth of Chlamydomonas reinhardtii for commercial production

Fields, Frank
UC San Diego

Chlamydomonas reinhardtii has long been a model organism for understanding algal genetics and exploring the production of transgenic protein. Here we show the potential of this organism to be more than a model, but a successful production platform for commercial use. 11 common lab strains of CR were initially screened to identify a high-productivity strain. The wild type of CC2937 was chosen and grown mixotrophically in a pH-auxostat batch-fed photobioreactor with acetic acid as the main carbon source. Over the course of 10 days, cell densities peaked at 8.7x10^7 cells/mL and biomass reached a high of 18.5 g/L (AFDW).

Closing the Loop - All Under One Roof

Roger's Community Garden
UC San Diego

At Roger's Community Garden's our goal is to unite a wide variety of disciplines including art, science, engineering, and more in order to innovate and bring together new solutions for food, fuel, and water. This includes anaerobic digestion, which has dual-benefits by redirecting food-waste from land-fills by turning it into bio-gas energy. Co-opting nature in the form of worms and larvae to consume food-waste also creates two solutions by again removing food/animal waste and creating fish feed for aquaponics. Aquaponics being the utilization of a closed-loop form of growing fish with food (and bacteria!) hydroponically to use only 10% of the water of traditional agriculture. Atmospheric water generation is then used powered via these renewable energies to condense water from the air to allow for water-neutral protein, vegetable, and fruit production (or even positive for potable drinking water). All of these technologies coming together to provide an off-grid solution for the food, energy, and water trilema that faces the world by not only mitigating climate change, but also adapting to it as well.

Genome and methylome of the oleaginous diatom Cyclotella cryptica reveal genetic flexibility toward a high lipid phenotype

Traller, Jesse
Scripps Institution of Oceanography, UC San Diego

Cyclotella cryptica is a coastal marine diatom that was previously studied during the US DOE’s Aquatic Species Program and deemed a strain suitable for large-scale biofuel production. Today, with a myriad of advancements and techniques in high throughput sequencing, it is becoming feasible to obtain high quality, cost-efficient genomic data on potential production strains, such as C. cryptica. We sequenced the genome, methylome, transcriptome, and proteome of C. cryptica and used comparative genomics to elucidate carbon metabolism and gene regulation in the cell. In particular, our efforts were focused on annotation of genes putatively involved in regulation of terminal carbon-rich pools, such as triacylglyceride, chrysolaminarin, and chitin. Results from this study highlight genes, genomic and epigenetic features, and metabolic processes that are of interest from an industrial production perspective as well as enhance our understanding of algal cellular biology.

 

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