Source: UNIVERSITY OF CALIFORNIA, DAVIS submitted to NRP
UNDERSTANDING ENDOMEMBRANE TRAFFICKING, CELL WALL BIOSYNTHESIS AND DEPOSITION IN PLANTS FOR CROP IMPROVEMENT, BIOFUEL GENERATION AND FOOD
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
1008039
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Oct 2, 2015
Project End Date
Sep 30, 2020
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
Plant Sciences
Non Technical Summary
The endomembrane system is responsible for the synthesis, delivery and maintenance of some of the most biologically interesting cellular products in plants, including cell wall components, active enzymes, products utilized in plant defense to pathogens and storage proteins.Plant cell walls offer a potentially limitless source of renewable energy, and a greater understanding of cell wall synthesis and assembly could facilitate the development of more accessible biofuels. Although the structure of cell walls is generally understood and the synthesis of cell wall components is characterized, remarkably little is known about how polysaccharides and the cellular machinery essential for cell wall assembly are transported through the cell to the apoplast. Components of the cell wall are thought to be transported from their synthesis sites to the plasma membrane through the endomembrane system, using a largely unknown mechanism.The proposed research will address questions concerning how plant cells direct different polysaccharides and components of cell wall assembly "machinery" to the sites where polysaccharides are deposited into the cell wall. The dissection of this complex system will be achieved through a comprehensive analysis of its interacting structural and functional components using complementary approaches: chemical genomics and classical genetics. As a longer term goal we would like to identify vesicles and components in the pathways that are essential for storage of cereal seed proteins or responses to environmental factors including pathogens. This knowledge could lead in the modification of crop plants towards increased tolerance to biotic and abiotic stress, increased biomass production and more accessible biofuels.
Animal Health Component
25%
Research Effort Categories
Basic
75%
Applied
25%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2012499108010%
2012499103010%
2032499103010%
2032499100010%
2032499104010%
2062499100010%
2062499103010%
2062499104010%
2062499108010%
2032499102010%
Knowledge Area
206 - Basic Plant Biology; 203 - Plant Biological Efficiency and Abiotic Stresses Affecting Plants; 201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
2499 - Plant research, general;

Field Of Science
1040 - Molecular biology; 1030 - Cellular biology; 1000 - Biochemistry and biophysics; 1080 - Genetics; 1020 - Physiology;
Goals / Objectives
GENERAL GOAL:Future economic growth and prosperity will depend on the availability of sustainable clean energy resources that protect the environment and reduce our dependence on fossil fuels. Plants provide one of the most promising sources of clean energy because they use sunlight to fix atmospheric carbon, producing fermentable sugars that can subsequently be converted into ethanol. There is a great potential in California towards utilizing advanced biofuel crops that could lead to cellulosic ethanol and/or other biofuels. This in turn could provide a significant part to the state energy needs. The overarching long-term goal of this multidisciplinary project is to investigate how plant cells direct different polysaccharides, cell-wall biosynthetic enzymes and other cell wall components into the developing cell wall. Plant cell walls are dynamic structures that play a pivotal role in growth and development, the maintenance of cell integrity and protection against pathogens. They are composed primarily of structural carbohydrates, and these can be broken down into sugars that provide energy directly, in food and feed, or indirectly by fermentation into ethanol. Plant cell walls therefore represent a potentially limitless source of renewable energy and a greater understanding of their synthesis and assembly could eventually lead to the development of more nutritious food/feed and more accessible biofuels and promising strategies for increasing stress tolerance. However, much of this fixed carbon is found as structural carbohydrates such as the cellulose in plant cell walls, and the inefficiency of the conversion of cellulose and other polysaccharides back into fermentable sugars is one of the major bottlenecks in the development of a sustainable biofuel industry (Pauly and Keegstra, 2008; Liepman et al., 2010). Unlocking the potential of plant cell walls will require more investigation of their structure and composition, greater understanding of the synthesis and assembly of structural carbohydrates in the context of the plant cell as a complex functional network, and novel strategies to modify the underlying processes, making the production of bioethanol and other biofuels more inexpensive and efficient. Although the structure of cell walls is understood in some detail and the synthesis of cell wall components is well characterized, remarkably little is known about the transport of these components to the plasma membrane. Polysaccharides and the cellular machinery required for cell wall assembly are thought to be transported from their sites of synthesis to the plasma membrane through a complex network of interconnected vesicles known as the endomembrane system, however this mechanism is largely unknown (Worden et al., 2012).JUSTIFICATION: Understanding this transport process would allow the introduction of modifications facilitating the conversion of plant biomass into fuel or increase the nutritional value of plants that are part of the human diet. It would also provide valuable information about plant growth and development, and interactions between plants and the environment, including pathogens.The objective of this multidisciplinary project is to investigate the role of endomembrane system in cell wall deposition, plant development and response to environmental stimuli. The dissection of this complex system will be achieved through the comprehensive analysis of its interacting structural and functional components using several complementary approaches (classical genetics and chemical genomics and vesicle isolation), and by developing novel strategies to characterize cell wall deposition. Extending this line of research this project will be further exploring the role of endomembrane system in crop improvement and food production. Regulatory mechanisms of the endomembrane trafficking will be further tested for their effect in trafficking important molecules in food production, pathogen and abiotic stress response. Both monocot and dicot plants will be used in this studies. Arabidopsis and algae will be used as a screening tool to identify new genes and chemicals that will be further explored in other plant species. A novel approach of chemical genomics will used to identify chemicals that can be used to dissect endomembrane trafficking mechanisms that control polysaccharide deposition and assembly and responses of plants to external stimuli. One of the advantages of chemical genomics is translatability, thus selected chemicals can be used to identify similar mechanisms in several plant species including crop plants such as rice wheat and tomato.OBJECTIVES:1. To identify chemicals affecting endomembrane trafficking, cell wall biosynthesis, deposition and/or digestibility and can be used for economical strategies for cell wall modification. 2. To identify genes involved in cell wall biosynthesis, transport, deposition and assembly of polysaccharides.3. To identify components of endomembrane trafficking that are involved in responses of plants to environmental signals and can be used for crop improvement.4. To train undergraduate and graduate students and postdoctoral scholars in molecular and cellular biology, classical genetics and chemical genomics and in critical thinking.Literature Cited:Liepman AH, Wightman R, Geshi N, Turner SR, Scheller HV (2010) Arabidopsis - a powerful model system for plant cell wall research. Plant Journal 61: 1107-1121Pauly M, Keegstra K (2008) Cell-wall carbohydrates and their modification as a resource for biofuels. Plant J 54: 559-568Worden N, Park E, Drakakaki G. (2012) Trans-Golgi network: an intersection of trafficking cell wall components. J Integr. Plant Biol. 54(11):875-86
Project Methods
Objective 1 and 2:Chemical genomic approaches to identify probes that affect cell wall biosynthesis and deposition. We will perform a chemical genomics screen to identify chemical probes that regulate endomembrane trafficking and the deposition of cell wall polysaccharides. The identified inhibitors will provide useful additions to a toolkit that can be used by the trafficking/cell wall research community. Chemical genomics has been used to modify the activity of specific proteins and pathways rapidly, reversibly and conditionally, complementing genetic screens and overcoming difficulties caused by gene lethality and redundancy (Drakakaki et al., 2009). A sub-library of 360 chemicals that affect endomembrane trafficking was identified in a screen of approximately 48,000 compounds carried out by the PI (Drakakaki et al., 2011). Using this approach we have identified novel probes that affect callose deposition during cytokinesis (Park et al., 2014; Davis et al., 2015) or cellulose deposition during cell elongation (Worden et al., 2015). Characterization of their mechanism of action, though the identification of their target pathways will identify new components involved in the deposition of callose and or cellulose. Genetic screens will be performed to identify molecular players in the pathways targeted by the small molecules. A whole genome sequencing approach will be used to identify the causal genes in the resistant mutants. Identification of the candidate genes and characterization of their function will provide insights in the regulation of these polysaccharides. To aid mutant characterization polysaccharide analysis will be performed using standard approaches wall analytical techniques. The Metabolomics Facility in the UC Davis Genome Center will facilitate use of the oligosaccharide mass profiling technique developed by Markus Pauly et al. (Obel et al., 2009). In a complementary approach, analysis of cell walls will be performed in situ utilizing the substantial array of wall component-specific monoclonal antibodies (mAbs) available at the CCRC (Pattathil et al., 2010). This methodology will enable us to identify with some degree of specificity, local changes in wall composition that result from tissue exposure to small molecules.Additional small molecules derived from the screens described above will be characterized using the same approach. If small molecules will cause specific vesicles to be mislocalized under chemical treatment, a vesicle isolation methodology (Drakakaki et al., 2012) will be used to prepare vesicles affected by the small molecules which will then be subjected to parallel proteomic and polysaccharide analysis to identify the protein and polysaccharide cargo.Genetic screens to identify new players in endomembrane trafficking and cell wall formation.We will carry out genetic screens to identify mutants that affect endomembrane trafficking and cell wall deposition. A mutagenized population of Arabidopsis plants expressing a trans-Golgi marker protein will be screened further for endomembrane trafficking and cell wall defects. Characterization of these mutants and identification of the causal genes will be performed as described above. Characterization will include polysaccharide composition analysis, subcellular characterization and morphological analysis. Once genes are identified they can be tested in crop plants as well. Objective 3: The approaches used above will lead to the identification of several gene products that are important in trafficking of cell wall components. These can include either proteins involved in endomembrane trafficking contributing to endomembrane structure and integrity or proteins involved in cell wall biosynthesis, deposition and assembly. The participation of these proteins in responses of plants to environmental signals and their use for crop improvement will be examined. Towards these goals, tools will be developed in crop plants such as Pistachio or Almond to determine abiotic and biotic stress responses. Those will include imaging the subcellular localization of sodium in different rootstocks in order to dissect mechanisms that confer salt tolerance. Literature Cited:Davis DJ, McDowell SC, Park E, Hicks G, Wilkop TE, Drakakaki G (2015). The RAB GTPase RABA1e localizes to the cell plate and shows distinct subcellular behavior from RABA2a under Endosidin 7 treatment. Plant Signal Behav.10(3):e984520. doi: 10.4161/15592324.2014.984520.Drakakaki G, Robert S, Raikhel NV, Hicks GR (2009) Chemical dissection of endosomal pathways. Plant Signal Behav 4: 57-62Drakakaki G , Robert S, Szatmari A-M, Brown MQ, Nagawa S, Van Damme D, Leonard M , Yang Z, Girke T, Schmid SL, Russinova E, Friml Jb, Raikhel NV, and Hicks GR. (2011) Clusters of bioactive compounds target dynamic endomembrane networks in vivo. PNAS 108(43):17850-5. Drakakaki G, van de Ven W, Pan S, Miao Y, Wang J, Keinath N, Weatherly B, Jiang L, Schumacher K, Hicks G, Raikhel N (2012) Isolation and proteomic analysis of the SYP61 compartment reveal its role in exocytic trafficking in Arabidopsis. Cell Research. 2012 Feb;22(2):413-24. Obel N, Erben V, Schwarz T, Kuhnel S, Fodor A, Pauly M (2009) Microanalysis of plant cell wall polysaccharides. Mol Plant 2: 922-932Park E, Diaz-Moreno S, Davis D, Wilkop TE, Bulone V, and Drakakaki G.(2014). Endosidin 7 specifically arrests late cytokinesis and inhibits callose biosynthesis revealing distinct trafficking events during cell plate maturation. Plant Physiology doi: http://dx.doi.org/10.1104/pp.114.241497Pattathil S, Avci U, Baldwin D, Swennes AG, McGill JA, Popper Z, Bootten T, Albert A, Davis RH, Chennareddy C, Dong R, O'Shea B, Rossi R, Leoff C, Freshour G, Narra R, O'Neil M, York WS, Hahn MG (2010) A comprehensive toolkit of plant cell wall glycan-directed monoclonal antibodies. Plant Physiol 153: 514-525Worden N, Wilkop TE, Esteve VE, Jeannotte R, Lathe R, Vernhettes S, Weimer B, Hicks G, Alonso J, Labavitch J, Persson S, Ehrhardt D, Drakakaki G (2015). CESA TRAFFICKING INHIBITOR inhibits cellulose deposition and interferes with the trafficking of cellulose synthase complexes and their associated proteins KORRIGAN1 and POM2/CELLULOSE SYNTHASE INTERACTIVE PROTEIN1. Plant Physiol. 2015 Feb;167(2):381-93

Progress 10/02/15 to 09/30/20

Outputs
Target Audience:The target audience for this project includes growers, farm advisors, industry representatives, scientists and students in industry, commodity boards nationally and internationally and at UC Davis and off campus. The results were also presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards, farmers and farm advisors in CA. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences, Pistachio day and annual pistachio workshops, Pomology (PECC) PT Meetings) and state wide meetings in 2015-2020. Participants (industry representatives, scientists, students) at the American Society of Plant Biologists Meetings, San Jose, CA USA, at the MWPCD meeting Penn State University, USA, The ASCB Meeting San Diego CA, Scientists students at the University of KY Lexington, the Indiana University Bloomington, the Texas A&M University, College Station, the University of Purdue, Ohio university, the Joint International Symposium of BAICFTBMD and UPSC, Beijing China, the Cold Spring Harbor -Asia Conference, Gyeongju, Korea, the 2016, 2017 and 2018 Cell Wall Meetings, the ENPER meeting, Prague Czech Republic, the Greek Agricultural Organization (ELGO Dimitra), the Mediterranean Agronomic Institute of Chania, Greece, the Hebei Agricultural University and the Zhejiang A&F University in China and the Norwegian University of Science and Technology. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Within this project the following individuals were involved: Graduate student Destiny Davis, Natasha Worden, Rosalie Sinclair and Shuxiao Zhang. The postdoctoral researchers Dr. Michel Ruiz Rosquete and Mingqin Chang, the undergraduate students: Karen Nguyen, Oliver Betz, Akaash Prasad, Fiona Dale-Huang, Maximilian Probstel, Angelica Tujo, Emily Mabry, Neije Mukherjee-Roy and the international visiting scholars: Guangxi Ren, Yuhang Shao, Hongguang Pang, Wenlong Bao, Yukun Cheng, Shahab Madahhosseini, Tereza Ticha, Hou-Ling Wang, Sina Pfleger, Alessandra Quartararo and the high-school students Phoebe Loo, Yolanda Shen, Connor Melendrez, Sean Fu and John Almazan. How have the results been disseminated to communities of interest?We have disseminated our results in the form of publications, reports and scientific presentations at meetings and workshops at national and international level. Presentations and reports were presented to growers, farm advisors, industry representatives, scientists and students in industry, commodity boards nationally and internationally and at UC Davis and off campus. Further, the results were also presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards, farmers and farm advisors in CA. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences, Pistachio day and annual pistachio workshops, Pomology (PECC) PT Meetings) and state wide meetings in 2015-2020. Participants (industry representatives, scientists, students) at the American Society of Plant Biologists Meetings, San Jose, CA USA, at the MWPCD meeting Penn State University, USA, The ASCB Meeting San Diego CA, Scientists students at the University of KY Lexington, the Indiana University Bloomington, the Texas A&M University, College Station, the University of Purdue, Ohio university, the Joint International Symposium of BAICFTBMD and UPSC, Beijing China, the Cold Spring Harbor -Asia Conference, Gyeongju, Korea, the 2016, 2017 and 2018 Cell Wall Meetings, the ENPER meeting, Prague Czech Republic, the Greek Agricultural Organization (ELGO Dimitra), the Mediterranean Agronomic Institute of Chania, Greece, the Hebei Agricultural University and the Zhejiang A&F University in China and the Norwegian University of Science and Technology. What do you plan to do during the next reporting period to accomplish the goals?We will continue our efforts towards dissecting cell wall deposition and understanding plant responses to biotic and abiotic stresses.

Impacts
What was accomplished under these goals? The multidisciplinary research project has at its center the dissection of endomembrane pathways directing cell wall components during plant growth, development and stress responses. Plant cell walls are a dynamic network of polysaccharides, proteins and other polymers essential for plant growth, while also providing a natural barrier against pathogens and a defense layer against abiotic stresses. Cell wall polysaccharides are valuable food sources, as dietary fibers are important in reducing the risk of contracting serious human diseases, while they are also useful as sources for feed, general fiber material and biomass that can be converted to biofuel. Although the structure of plant cell walls is generally understood, remarkably little is known about the transport mechanisms of polysaccharides and the cell wall biosynthesis machinery. This transport is mediated through a complex network of subcellular compartments called the endomembrane system. Understanding mechanisms of cell wall build-up can form frameworks for strategies modifying plant characteristics towards nutritional improvement and enhanced responses under biotic and abiotic stresses and biomass utilization. Uncovering mechanisms of cell wall build-up Uncovering mechanisms of cell wall build-up can help to develop strategies improving desired cell wall characteristics for agriculture and human health. Endomembrane pathways tightly regulate cell wall development, however these mechanisms are largely unknown. Knowledge of the type of carbohydrate cargo delivered by transport compartments, known as vesicles, containing building blocks of cell wall, is essential for a comprehensive understanding of plant plasticity under different conditions. Pioneering a method of plant vesicle isolation coupled with large-scale polysaccharide detection allowed us to identify, for the first time, the glycan contents of specific subsets of transport vesicles. This study gave an unprecedented view of the structure and composition of wall polysaccharides still within the endomembrane system, details that have been unknown for decades. Our data point to a secretion of "polysaccharide building blocks" prior to their assembly in the cell wall, challenging the current dogma. Importantly, this prompts us to investigate how these glycans, while in the endomembrane system, change during plant development or as a response to environmental stimuli and stresses, a direction that my lab is currently focusing on. With this knowledge, we can design strategies for desirable cell wall characteristics towards improved nutritional value, biomass production, resistance to pathogens and ability to cope with climate change. Regulation of cellular pathways involved in stress response Uncovering how are cellular transport pathways regulate and control stress response, such as salinity, can help us develop tolerance mechanisms and select superior genotypes for breeding and cultivation. Transport between membrane compartments in a cell is a finely regulated process to ensure correct cargo delivery and proper plant cell function. We identified a novel regulator of cellular trafficking, namely the tethering complex TRAPPIII and studied one of its subunits, AtTRAPPC11, in detail. This evolutionarily conserved subunit plays a role in maintaining the organization of cellular compartments that regulate secretion, likely acting as a checkpoint mechanism of transport control. Importantly, this novel protein in plants is important for plant response to salinity stress, an undescribed role of plant tethers in abiotic stress. Given that human TRAPC11 mutations are responsible for a number of human diseases, we similarly expect to uncover how this complex as a whole or its specific subunits control plant growth and environmental biotic and abiotic stress response. Dissecting salinity response in almond and pistachio rootstocks Pistachio and Almonds are two of the most economically valuable crops in California with acreage expanding at a great rate. Development of rootstocks adapted to California's other main cropping challenge, changing climate, is important for crop sustainability and economic viability. So far, most pistachio and almond rootstock characterization is based on surveying the aerial part (above ground organs, leaves and shoots). However, it remains challenging to select for superior rootstocks without taking into account the root system, and how it adapts to environmental stresses. Plant roots are the first contact to nutrients and microorganisms and are important for robust and sustainable growth. Bidirectional cell barriers with reinforced cell walls such as the endodermis surrounding the vascular tissue, control access to the root conducting tissues and prevent solute leakage. We are studying how cell wall deposition and cell barrier differentiation is altered under abiotic stress, during which impermeable cell wall polymers are deposited, controlling the transport of toxic ions. Toward this end, we developed methodologies for the detection of the implicated ions in salinity stress at the subcellular level in live tissues and combined it with anatomical characterization across a root developmental gradient. Our combination of laboratory and green house studies showed that: I) sodium sequestration in the vacuole of root cortex cells in combination with ion exclusion is associated with salt tolerant seedlings. II) a root developmental gradient controls root salinity response. Further, reinforcement of cellular barriers (endodermis and exodermis) by suberin deposition correlates with salinity tolerance. Overall, foundational studies revealed mechanisms of salinity tolerance in almond and pistachio rootstocks from cellular and structural perspectives across a root developmental gradient and provide insights for future screens targeting stress response in tree nut crops.

Publications

  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Ren G, Ruiz Rosquete M, Peralta AG, Pattathil S, Hahn MG, Wilkop T, Drakakaki G (2020). Isolation and Glycomic Analysis of Trans-Golgi Network Vesicles in Plants. Methods Mol Biol., 2177: 153-167.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Rosquete MR, Worden N, Drakakaki G (2019). AtTRAPPC11 is involved in TRAPPIII mediated control of post-Golgi protein trafficking. Plant Signal Behav. 14 (12):1676631. doi: 10.1080/15592324.2019.1676631.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Rao X, Bartley LE, Drakakaki G, Anderson CT. Regulation of and by the Plant Cell Wall (2020). Front Plant Sci. Apr 29;11:513. doi: 10.3389/fpls.2020.00513.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Davis DJ, Wang M, S�renson I, Rose JKC, Domozych D, Drakakaki G (2020). Callose deposition is essential for the completion of cytokinesis in the unicellular alga, Penium margaritaceum. Journal of Cell Science, Oct 12;133(19):jcs249599.
  • Type: Other Status: Published Year Published: 2020 Citation: Pang H, Shao Y, Wang M, He F, Madahhosseini S, Zhang S, Betz O, Jernstedt J, Espinoza M, Wilkop T, Drakakaki G. Molecular and subcellular characterization of salinity tolerance in almond rootstocks. Annual Report, Almond Research Board. 2020
  • Type: Other Status: Published Year Published: 2020 Citation: Shao Y, Pang H, Wang M, He F, Madahhosseini M, Zhang S , Betz O, Jernstedt J, Espinoza M, Wilkop T. Drakakaki G. Characterization of root anatomy and plasticity in almond rootstocks for improved nutrient uptake and stress response. Annual Report, Almond Research Board. 2020
  • Type: Other Status: Published Year Published: 2020 Citation: Zhang S, Betz O, Quartararo A Madahhosseini S, Wilkop T, Judy Jernstedt, Michelmore R, Ferguson L, Drakakaki G. Characterization of root plasticity in pistachio rootstocks for better nutrient uptake and stress response. Annual Report, Pistachio Research Board. 2019
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Shao Y, Pang H, Wang M, He F, Madahhosseini S, Espinoza M, Zhang S,Betz O, Wilkop T, Jernstedt J, Drakakaki G. Morphological and Cellular Characterization of Salinity Tolerance in Different Genotypes of Almond Rootstocks., The Almond Conference. December 10-12, 2019 Sacramento, California. Poster.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Pang H, Shao Y, Wang M, He F, Madahhosseini S, Espinoza M, Zhang S, Betz O, Wilkop T, Jernstedt J, Drakakaki G. Uncovering Root Plasticity and Cellular Responses of Almond Rootstocks under Salinity Stress, The Almond Conference. December 10-12, 2019 Sacramento CA. Poster.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Drakakaki G. Root plasticity under salinity stress in rootstocks and approaches to dissect pistachio fruit development, Pistachio research board meeting. UC Merced Center. 2020 January 28.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Drakakaki G. Interplay between endomembrane dynamics and cell wall formation, Department of Botany & Plant Pathology, Purdue University, March 19, 2020
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sinclair R, Davis D, Jawaid Z, Cox D, G Drakakaki. Cytokinesis and the importance of Callose. UC Davis Plant Biology Retreat, UC Davis, CA. October 26th, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Davis, D.J., Domozych, D.S., Drakakaki, G. Callose is essential for cytokinesis in the unicellular Charophyte, P. margaritaceum. Plant Biology Retreat, October 26th, 2019. Davis, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Jawaid MZ., Drakakaki G., Cox D. A Biophysical Model of Intermediate Phases in Developing Cell Plates. UC Davis Plant Biology Retreat, UC Davis, CA. October 26th, 2019.


Progress 10/01/18 to 09/30/19

Outputs
Target Audience:The target audience for this project includes growers, farm advisors, industry representatives, scientists and students in industry, commodity boards nationally and internationally and at UC Davis and off campus. The results were presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences (Pistachio day and annual pistachio workshop) and state meetings in 2018-2019. Participants (industry representatives, scientists, students) at the American Society of Plant Biologists Meeting, San Jose, CA USA, at the MWPCD State College, USA, The ASCB Meeting San Diego CA, Scientists students at University of KY Lexington, and at the Norwegian University of Science and Technology Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Within this project the following individuals were involved: Graduate student Destiny Davis, Rosalie Sinclair and Shuxiao Zhang. The postdoctoral researcher Dr. Michel Ruiz Rosquete, the undergraduate students: Oliver Betz, Akaash Prasad, Fiona Dale-Huang and the international visiting scholars: Guangxi Ren, Yuhang Shao, Hongguang Pang, Wenlong Bao, Yukun Cheng, Shahab Madahhosseini and the high-school student Phoebe Loo. How have the results been disseminated to communities of interest?We have disseminated our results in the form of publications, reports and scientific presentations at meetings and workshops at national and international level. Presentations and reports were presented to commodity boards, industry representatives and farm advisors. Further, the results were presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences (Pistachio day and annual pistachio workshop) and state meetings in 2018-2019. Participants (industry representatives, scientists, students) at the American Society of Plant Biologists Meeting San Jose, CA USA, at the MWPCD meeting, State College USA, The ASCB meeting San Diego CA, Scientists and students at University of KY Lexington, and at the Norwegian University of Science and Technology. What do you plan to do during the next reporting period to accomplish the goals?We will continue our efforts towards dissecting cell wall deposition and understanding plant responses to biotic and abiotic stresses.

Impacts
What was accomplished under these goals? Plants have developed an unusual level of developmental plasticity compensating for their sessile lifestyle, an adaptive behavior noticeable throughout their entire life cycle. Adaptations include plant specific compartments such as the cell wall, a complex macromolecular composite polysaccharide structure surrounding and protecting plant cells, essential for plant growth, development, signal transduction, and disease resistance. Vesicle trafficking pathways control the tightly regulated cell wall development. However, little is known about the mechanisms that orchestrate endomembrane-based transport of cell wall components that control cell wall development. This is partly due to the significant challenges posed by isolating endomembrane compartments and determining their polysaccharide cargo. Towards this end, we conduct highly multidisciplinary research, that has at its center the development of innovative methodologies for the dissection of mechanisms involved cell wall formation in plant growth, and stress responses. 1) We pioneered a method of plant vesicle isolation that allowed us to separate post Golgi vesicles displaying the trans-Golgi Network (TGN) SNARE, Syntaxin of Plants 61, SYP61 and analyzed their proteome. This analysis identified novel players in the endomembrane system and suggested that the SYP61 pathway is involved in cell wall deposition and plant stress responses. Building on this knowledge, we recently developed the glycome profile of SYP61 vesicles by combining vesicle isolation with large-scale carbohydrate antibody arraying. This study gave an unprecedented view of the structure of wall polysaccharides still within the endomembrane system, details which have been unknown for decades. Diverse glycan structures of both pectin xyloglucan and glycoprotein epitopes were identified in the isolated vesicles, indicating a common pathway for both types of polysaccharides. Follow-up studies included the interrogation of xyloglucan- and pectin-specific epitopes in the cell wall of the Arabidopsis SYP61 mutant via in muro and glycome assays, demonstrating the role of the SYP61 pathway in cell wall development (Wilkop et al., 2019). Overall, our study provides the first large scale array of TGN compartments involved in transport and deposition of structural polysaccharides, answering long term standing questions in the field. We identified the tethering complex TRAPPIII and studied one of its subunits, AtTRAPPC11, in detail. Our study revealed that this evolutionarily conserved subunit plays a role in maintaining the organization of the TGN and endosomal trafficking, likely acting as a checkpoint control mechanism at the TGN. In addition, AtTRAPPC11 is important for the plant response to salinity stress, an undescribed role of these proteins in abiotic stress (Rosquete et al., 2019). Identification and further characterization of this complex opens new avenues to explore the role of tethering factors in plant environmental responses. 2) Pistachio and almond is a rapidly expanding commodity in California. With the decreasing availability of fresh water, increasing climate instability which requires irrigation, and thus increasing problems with soil quality, pistachio has become an ideal candidate for production in adverse conditions. Development of rootstocks that adapt to California's changing climate is important for crop sustainability and optimal growth. So far, most of the rootstock characterization is based on aerial part (above ground organs, leaves and shoots). Plant roots are the first contact to nutrients, and microorganisms and are important for robust and sustainable growth. However, it remains challenging to select for superior rootstocks without taking into account their root system, and how it adapts to environmental stresses. Our objective is to identify, physiological, cellular, anatomical and molecular determinants that contribute to salinity tolerance and root adaptation in pistachio and almond rootstocks. In depth analysis of pistachio and almond rootstocks using our developed methodologies for ion detection and root morphological analysis cumulative show that: A root developmental gradient controls root response. The major activity with respect to ion sequestration occurs in the very tip of roots. Control of ion uptake and redistribution contributes to salinity tolerance. Increased suberin deposition in cellular barriers (endodermis and exodermis) is correlated with salinity tolerance. Based on our results we started the development of molecular markers for salt and abiotic stress tolerance screening of pistachio and almond rootstock populations. Overall, our efforts can assist in the characterization of complex mechanisms that control salinity stress of woody plants.

Publications

  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Wilkop T, Pattathil S, Ren G, Davis DJ, Bao W, Duan D, Peralta AG, Domozych D, Hahn MG, Drakakaki G. (2019). A hybrid approach enabling large scale glycome analysis of post-Golgi vesicles reveals a transport route for polysaccharides. Plant Cell. Mar;31(3):627-644 (Faculty of 1000 recommended).
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Rosquete MR, Worden N, Ren G, Sinclair R, Pfleger S, Salemi M, Phinney B, Domozych D, Wilkop T, Drakakaki G. (2019). AtTRAPPC11/ROG2: a role for TRAPPs in maintenance of plant TGN/EE organization and function. Plant Cell. Aug;31(8):1879-1898.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Wang M, Zhang L, Boo KH, Park E, Drakakaki G, Zakharov F. (2019). PDC1, a pyruvate/?-ketoacid decarboxylase, is involved in acetaldehyde, propanal and pentanal biosynthesis in melon (Cucumis melo L.) fruit. Plant J. Apr;98(1):112-125.
  • Type: Other Status: Published Year Published: 2018 Citation: Drakakaki G Subcellular and molecular characterization of salinity tolerance in almonds with novel tools 2018 Co-operative personnel: Oliver Betz, Shahab Madahhosseini, Fiona Dale-Huang, Malli Aradhya, John Preece, Bruce Lampinen, Patrick Brown, Tom Gradziel, Roger Duncan, John Labavitch, Judy Jernstedt, Thomas Wilkop. Annual Report, Almond Research Board. 2018
  • Type: Other Status: Published Year Published: 2019 Citation: Drakakaki G, Zhang S, Betz O, Quartararo A Madahhosseini S, Wilkop T, Judy Jernstedt, Michelmore R, Ferguson L. Characterization of root plasticity in pistachio rootstocks for better nutrient uptake and stress response. Annual Report, Pistachio Research Board. 2018
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Madahhosseini S , Betz O, Wilkop T, Dykes B, Jernstedt J, Drakakaki G. Subcellular Characterization of Salinity Tolerance in Almond Rootstocks Annual Almond Conference. Sacramento CA, USA. December 4- 6, 2018.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Drakakaki G. Characterization of root plasticity in pistachio rootstocks for better nutrient uptake and stress response. Pistachio Meeting, Visalia CA. January 17, 2019. Oral Presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Drakakaki G. Subcellular Characterization of Salinity Tolerance in Almond Rootstocks Annual Almond Conference. Sacramento CA, USA. December 4-6, 2018. Oral Presentation
  • Type: Other Status: Published Year Published: 2019 Citation: Drakakaki G. Cellular dynamics and cell wall development. Norwegian University of Science and Technology, Trondheim Norway , September 15, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Drakakaki G. Endomembrane dynamics during plant cell division. Plant Cell Dynamics Meeting, State College, PA. 2019 June 18-21.
  • Type: Other Status: Published Year Published: 2019 Citation: Drakakaki G. Plant endomembrane dynamics and glycomics. University of Kentucky Lexington. March 13, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. Dissecting endomembrane dynamics. Annual Plant Biology Retreat, Tahoe CA. October, 20, 2018.
  • Type: Other Status: Published Year Published: 2018 Citation: Drakakaki G. Polysaccharide deposition during cytokinesis. UC Davis Fall Seminars. UC Davis, 10 October 2018.
  • Type: Other Status: Published Year Published: 2019 Citation: Sinclair R M. Towards modeling cell plate dynamics". Oral presentation. Plant Biology Graduate Group, Progress Seminar, May 7, 2019.
  • Type: Other Status: Published Year Published: 2019 Citation: Rosquete M. A role for TRAPPs in maintenance of plant TGN/EE organization and function UC Davis Cell Biology Seminars. UC Davis, 10 May 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Davis, D.J., Domozych, D.S., Drakakaki, G. Cytokinesis, the Cell Wall, and a Charophyte. Plant Biology Retreat. October 21st, 2018. Tahoe City, CA. Contributed talk by Davis.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Rosquete MR, Worden N, Ren G, Sinclair R, Pfleger S, Salemi M, Phinney B, Domozych D, Wilkop T, Drakakaki G. AtTRAPPC11/ROG2: a role for TRAPPs in endomembrane trafficking and plant stress response. Annual Plant Biology Retreat, Tahoe CA. October, 20, 2018. Contributed talk by Rosquete.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: R. Sinclair, D. Davis Z. Jawaid, Jose Alonso, David Domozych, D. Cox, G. Drakakaki; Cytokinesis Through The Lenses Of Callose Plant Cell Dynamics Meeting, State College, PA. 2019 June 18-21. Contributed talk by Sinclair.
  • Type: Other Status: Published Year Published: 2018 Citation: Rosquete M. AtTRAPPC11/ROG2: a role for TRAPPs in maintenance of plant TGN/EE organization and function. UC Davis Plant Sciences Flash Fall Seminars. UC Davis, 10 October 2018.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Betz O*, Zhang S, Sheikhi A, Quartararo A, Madah-Hosseini S, Jernstedt J, Brown PJ, Drakakaki G. Characterization of root plasticity in pistachio rootstocks for better nutrient uptake and stress response Presented at the 30th Annual UC Davis Undergraduate Research Conference on April 26, 2019. *Undergraduate student presentation. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Prasad A*, Sinclair R, Drakakaki G. Dissecting the Role of Callose During Cell Plate Formation. Presented at the 30th Annual UC Davis Undergraduate Research Conference on April 26, 2019. *Undergraduate student presentation. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sinclair R. Plant Cell Wall Development: Modeling Live Cell Dynamics Oral Presentation. College outreach. Pine Mannor College, Chestnut Hill, MA. April 18 2019. Contributed talk by Sinclair.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Jawaid MZ., Drakakaki G., Cox D. A Biophysical Model of Intermediate Phases in Developing Cell Plates. APS March Meeting 2019 March 4 -8, 2019. Boston, Massachusetts. Contributed talk by Jawaid.


Progress 10/01/17 to 09/30/18

Outputs
Target Audience:The target audience for this project includes growers, farm advisors, industry representatives, scientists and students in industry, commodity boards nationally and internationally and at UC Davis and off campus. The results were presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences and state meetings. Participants (industry representatives, scientists, students) at the 2018 Cell Wall Meeting, CA USA, at the PAPMBW meeting Vancouver, Canada, the 42nd New Phytologist Symposium, CA USA, at the Joint International Symposium of BAICFTBMD and UPSC, Beijing China. Scientists, students and industry representatives the Greek Agricultural Organization (ELGO Dimitra), Chania, Greece, at the Indiana University Bloomington, the Agricultural University of Athens, Athens, Greece, the Texas A&M University, College Station, the Zhejiang A&F University Hangzhou, China and the UC Davis. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Within this project the following individuals were involved: Graduate student Destiny Davis and Rosalie Sinclair. The postdoctoral researcher Dr. Michel Ruiz Rosquete, the undergraduate students: Oliver Betz, Angelica Tujo, and the international visiting scholars: Tereza Ticha, Guangxi Ren, Wenlong Bao, Yukun Cheng, Jingrui Li, Shahab Madahhosseini and the high-school students: Connor Melendrez, Sean Fu and Yolanda Shen. How have the results been disseminated to communities of interest?We have disseminated our results in the form of publications, reports and scientific presentations at meetings and workshops at national and international level. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, farm advisors, industry representatives, scientists, students) at the Annual Almond and Pistachio conferences and state meetings. Participants (industry representatives, scientists, students) at the 2018 Cell Wall Meeting CA USA, the PAPMBW meeting in Vancouver, Canada, the 42nd New Phytologist Symposium, CA USA, the Joint International Symposium of BAICFTBMD and UPSC, Beijing China. Scientists and students at the Greek Agricultural Organization (ELGO Dimitra) Chania, Greece, the Agricultural University of Athens, Athens, Greece, the Texas A&M University, College Station, the Zhejiang A&F University Hangzhou, China, the Indiana University Bloomington and UC Davis. What do you plan to do during the next reporting period to accomplish the goals?We will continue our efforts towards dissecting cell wall deposition and understanding plant responses to biotic and abiotic stresses.

Impacts
What was accomplished under these goals? The plant cell wall is one of the evolutionarily conserved structures of terrestrial plants. The plant cell wall is essential for plant growth and response against abiotic stresses and pathogens. Our research is focused on investigating cellular mechanisms contributing to transport and deposition of cell wall polysaccharides and plant stress responses. 1) The plant endomembrane system facilitates the transport of polysaccharides, associated enzymes and glycoproteins through vesicle trafficking pathways controlling cell wall development. However, little is known about the mechanisms that orchestrate endomembrane-based transport of cell wall components and support plant growth and plant stress responses. This is partially attributed to technical challenges in biochemically determining polysaccharide cargo in specific vesicles. Addressing this limitation, we took a multidisciplinary approach to understand mechanisms contributing to the build up of cell wall and plant responses to biotic and abiotic stress. We developed a hybrid assay by combining vesicle isolation with large-scale carbohydrate antibody arraying, which enables charting the glycome profile of endomembrane compartments. We characterized post-Golgi vesicles marked by the SYP61 protein. Using a library of ~155 antibodies, recognizing specific non-cellulosic carbohydrate epitopes enabled us to identify a range of diverse glycans, including structural polysaccharides and glycoproteins in the isolated vesicles. Our study is the first of its kind, providing the first large scale array of trans-Golgi Network (TGN) compartments involved in transport and deposition of structural polysaccharides. It marks the beginning of a new frontier in the dissection of cellular pathways, that can be used to characterize cellular responses under different environmental stimuli, stresses, growth stages or disease conditions. 2) Pistachio and almonds are woody plants that are some of the major commercial crops in California. The current increase in global temperature in combination with intensive agriculture will substantially increase salt stress in crops, representing a major challenge for most of the irrigated cropland in California. Understanding the mechanism of salt ion uptake, transport, and sequestration at the cellular and molecular levels can provide a way to identify desirable plant characteristics that can be selected for, in rootstocks and scions in order to achieve optimal plant performance. Supported by the Almond and Pistachio Research Boards, we have developed the methodologies for the detection of the implicated ions in salinity stress, sodium, potassium and chloride. Our results are the first successful subcellular potassium and chloride imaging experiments in all plants. The plant root is a critical organ for nutrient acquisition that also forms an efficient defense against external stresses in order to maintain plant fitness. Bidirectional cell barriers such as the endodermis surrounding the vascular tissue, control the access to the root vasculature and prevent solute leakage. We investigate how cell barrier differentiation occurs under abiotic stress, during which impermeable cell wall polymers are deposited, strengthening the barrier zones and controlling the transport of toxic ions. In our pilot experiments, distinct, genotype specific, subcellular accumulation patterns of sodium were observed in rootstocks, demonstrating a likely exclusion mechanism in selected rootstock genotypes. We hypothesize that control of ion uptake and redistribution contributes to salinity tolerance; characterization of this mechanism is our current and future research focus. Our efforts are intended to assist in the characterization of complex mechanisms that control salinity stress of woody plants.

Publications

  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Rosquete M.R, Worden N, Wilkop T, Drakakaki G. "AtTRAPPC11/ROG2: a role for TRAPPs in maintenance of plant TGN/EE organization and function". 6th Pan American Plant Membrane Biology Workshop (PAPMBW). 24-28 June 2018. Vancouver, Canada. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. "Dissecting endomembrane dynamics and cell wall biogenesis" Joint International Symposium of BAICFTBMD and UPSC. Beijing China 27-30 August 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. "Endomembrane dynamics and polysaccharide deposition" 42nd New Phytologist Symposium. The biology of wood: from cell to trees. Lake Tahoe, USA18. - 22. June 2018.Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. Glycomic analysis enables characterization of vesicle traffic. VII Cell wall research conference, Asilomar, California, USA. 18.-22. June 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. ⿿Dissecting endomembrane dynamics and cell wall biogenesis⿝ Texas A&M University, College Station, TX. April 20, 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. "Endomembrane dynamics during cell division and plant morphogenesis". Indiana University Bloomington. February 15, 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. ⿿The building up cell wall⿝. Greek Agricultural Organization (ELGO Dimitra). Chania, Greece. October 16, 2017. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. ⿿Chemical biology to dissect endomembrane dynamics⿝. Agricultural University of Athens. Athens, Greece. October 9, 2017. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Davis D. ⿿Dissecting cytokinesis in Plant Cells with ES7⿝. Annual Plant Biology Retreat, 10-28-2017 Davis CA. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Rosquete M.R, Worden N, Wilkop T, Drakakaki G. "ROG2 encodes a putative TRAPPC11 subunit in Arabidopsis". Annual Plant Biology Retreat, 10-28-2017 Davis CA. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Davis, D.J., Park, E., McDowell, S., Comai, L., Drakakaki, G. (2018) Dissecting cell plate formation. Presented at UC Davis Plant Sciences Symposium. April 16, 2018, Davis, CA. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Betz O*, Quartararo A, Drakakaki G. Characterization of root plasticity in pistachio UCB-1 rootstocks for better nutrient uptake and stress response Presented at the 29th Annual UC Davis Undergraduate Research Conference on April 27, 2018. *Undergraduate student presentation. Poster
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Rosquete MR, Davis DJ, Drakakaki G. The Plant Trans-Golgi Network: Not Just a Matter of Distinction. Plant Physiol. 2018 Jan;176 (1):187-198
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Rosquete MR, Drakakaki G. Plant TGN in stress response: a compartmentalized overview. Current Opinion in Plant Biology. 2018:46 (1-8).
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Sinclair R, Rosquete MR and Drakakaki G. Post-Golgi trafficking and transport of cell wall components. Frontiers in Plant Science. doi: 10.3389/fpls.2018.01784.
  • Type: Journal Articles Status: Accepted Year Published: 2018 Citation: Wilkop TE, Pattathil S, Ren G, Davis D, Bao W, Duan D, Peralla AG, Domozych D, Hahn M, Drakakaki G. Glycomic analysis of vesicles reveals that SYP61 defines a secretory pathway regulating transport and deposition of structural polysaccharides. The Plant Cell. Accepted.
  • Type: Other Status: Published Year Published: 2018 Citation: Drakakaki G, Quartararo A, Ferguson L, Wilkop T (2018) Cellular, subcellular and molecular characterization of salinity tolerance in pistachio with novel tools. Annual Report. Pistachio Research Board.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. ⿿Characterization of salinity tolerance in pistachio rootstocks⿝. Oral Presentation , Pistachio Research Meeting. July 2018
  • Type: Other Status: Published Year Published: 2018 Citation: Drakakaki G ⿿Subcellular and molecular characterization of salinity tolerance in almonds with novel tools⿝ 2018 Co-operative personnel: Oliver Betz, Shahab Madahhosseini, Fiona Dale-Huang, Malli Aradhya, John Preece, Bruce Lampinen, Patrick Brown, Tom Gradziel, Roger Duncan, John Labavitch, Judy Jernstedt, Thomas Wilkop. Annual Report, Almond Research Board.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Cheng Y, Wilkop T, Quartararo A, Dorsey H, Jernstedt J, Drakakaki G. Subcellular Characterization of Salinity Tolerance in Almond Rootstocks Annual Almond Conference. Sacramento CA, USA. December 5-7, 2017. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Subcellular Characterization of Salinity Tolerance in Almond Rootstocks Annual Almond Conference. Sacramento CA, USA. December 5-7, 2017. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Drakakaki G. "Post Golgi pathways regulating plant growth". Zhejiang A&F University Hangzhou China 1st September 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Davis D, Wilkop T, Davis D, Pattathil S, Bao W, Peralta A, Duan, D, Domozych D, Hahn M, Drakakaki G. ⿿Packaged polysaccharides and the cellular pathways they take⿝. 6th Pan American Plant Membrane Biology Workshop (PAPMBW). 24-28 June 2018. Vancouver, Canada. Oral Presentation


Progress 10/01/16 to 09/30/17

Outputs
Target Audience:The target audience for this project includes industry, commodity boards nationally and internationally and scientists and students at UC Davis and off campus. The results were presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio Conferences. Participants (industry representatives, scientists, students) at the 2017 Cell Wall Meeting, the American Society of Plant Biologists (ASPB) meeting, the Midwest Plant Cell Dynamics Meeting, 20th European Network for Plant Endomembrane Research meeting. Scientists and students at University of Minnesota USA, Mediterranean Agronomic Institute of Chania, Greece and the Hebei Agricultural University, China. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Within this project the following individuals were involved: Project scientist: Thomas Wilkop. Graduate student Destiny Davis. The postdoctoral researcher Dr. Michel Ruiz Rosquete, the undergraduate students: Emily Mabry, Neije Mukherjee-Roy, Oliver Betz, and the international visiting scholars: Hou-Ling Wang, Wenlong Bao, Yukun Cheng, Sina Nicol Pfleger, Alessandra Quartararo, Longjun Chen, Jingrui Li and the high-school students: Jake Brugger and Yolanda Shen. How have the results been disseminated to communities of interest?We have disseminated our results in the form of publications, reports and scientific presentations at meetings and workshops and national and international level. Examples include: Members of the Almond and the Pistachio research boards. Participants (growers, industry representatives, scientists, students) at the Annual Almond and Pistachio conferences. Participants (industry representatives, scientists, students) at the 2018 Cell Wall Meeting Dalian China, the ASPB meeting, Hawaii , USA, the Midwest Plant Cell Dynamics Meeting, Madison USA, ENPER meeting, Prague Czech Republic, International Student Workshop. Nara Japan. Scientists and students at University of Minnesota USA, Mediterranean Agronomic Institute of Chania,Greece, Hebei Agricultural University, China and UC Davis. What do you plan to do during the next reporting period to accomplish the goals?We will continue our efforts towards dissecting cell wall deposition and understanding plant responses to biotic and abiotic stresses.

Impacts
What was accomplished under these goals? more info...) The plant cell wall is a network of polysaccharides and glycoproteins representing one of the evolutionarily conserved structures of terrestrial plants. The plant cell wall is essential for plant growth and response against abiotic stresses and pathogens. We investigate endomembrane trafficking mechanisms contributing to transport and deposition of cell wall polysaccharides and plant stress responses. Given the complexity of cell wall synthesis and assembly, we are using multidisciplinary approaches including chemical genomics, classical genetics, proteomics and vesicle glycomics and development ion staining methods to establish in detail the mechanisms contributing to the build up of a new cell wall and plant responses of plants to biotic and abiotic stress. 1) During the last two decades extensive studies of cell wall development have led to the identification of a number of enzymes involved in the biosynthesis of cell wall. However, comparatively little is known about their endomembrane-based transport, deposition and integration into the cell wall. The plant endomembrane system facilitates the transport of polysaccharides, associated enzymes and glycoproteins through the secretory pathway. Currently the nature of vesicles that carry cell wall components is not characterized, partly due to the inability to biochemically determine polysaccharide cargo in vesicles. By combining vesicle isolation with a large-scale carbohydrate antibody arraying technique we were for the first time able to characterize the glycome profile of trans-Golgi network (TGN) vesicles. We identified a range of diverse glycans, including pectins and xyloglucans in the vesicles, providing insights into the type of modification that individual cell wall macromolecules undergo en route to the wall or during the process of cell wall assembly. 2) Plant cytokinesis, a process fundamentally different from cytokinesis in animals and fungi, requires the de novo formation of new cell wall and partitions the cytoplasm of the dividing cell. We identified and characterized Endosidin 7 (ES7) as a specific inhibitor of callose deposition during cell division. We isolated and characterized mutants resistant to ES7, not exhibiting cell plate defects or a reduction of callose deposition at the cell plate upon ES7 exposure. Extending this genetic approach, we will identify new regulators of callose deposition and vesicle trafficking during cell division. Knowledge from this research impacts long-term biotechnological applications, including manipulating plant growth characteristics. 3) Pistachio and almonds are woody plants that are some of the major commercial crops in California. The current increase in global temperature in combination with intensive agriculture will substantially increase salt stress in crops. Soil salinization is a major challenge for most of the irrigated cropland in California. Understanding the mechanism of salt ion uptake, transport, and sequestration at the cellular and molecular levels can provide a way to identify desirable plant characteristics that can be selected for in rootstocks and scions in order to achieve optimal plant performance. Supported by the Almond and Pistachio Research Boards, we have developed the methodologies for the detection of the implicated ions in salinity stress, sodium, potassium and chloride. Our results are the first successful subcellular potassium and chloride imaging experiments in all plants. In our pilot experiments, distinct, genotype specific, subcellular accumulation patterns of sodium were observed in rootstocks, demonstrating a likely exclusion mechanism in selected rootstock genotypes. We hypothesize that control of ion uptake and redistribution contributes to salinity tolerance; characterization of this mechanism is our current and future research focus.

Publications

  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Smertenko A, Assaad F, Baluaka F, Bezanilla M, Buschmann H, Drakakaki G, Hauser MT, Janson M, Mineyuki Y, Moore I, M�ller S, Murata T, Otegui MS, Panteris E, Rasmussen C, Schmit AC, `amaj J, Samuels L, Staehelin LA, Van Damme D, Wasteneys G, }�rsk� V (2017) Plant Cytokinesis: Terminology for structures and processes. Trends Cell Biol. (17)30145-9
  • Type: Other Status: Published Year Published: 2017 Citation: Drakakaki G, Quartararo A, Ferguson L, Wilkop T. 2017. Cellular, subcellular and molecular characterization of salinity tolerance in pistachio with novel tools. Annual Report, Pistachio Research Board.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Characterization of salinity tolerance in pistachio rootstocks. Oral Presentation , Pistachio Research Board. January 2017
  • Type: Other Status: Published Year Published: 2017 Citation: Drakakaki G Subcellular and molecular characterization of salinity tolerance in almonds with novel tools Co-operative personnel: Cheng Y, Aradhya M, Preece J, Lampinen B, Brown P, Gradziel T, Duncan R, Dorsey H, Labavitch J, Wilkop T. Annual Report, Almond Research Board.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Cheng Y, Wilkop T, Quartararo A, Dorsey H, Jernstedt J, Georgia Drakakaki. Subcellular Characterization of Salinity Tolerance in Almond Rootstocks with Novel Tools. Annual Almond Conference. Sacramento CA, USA. December 6-8, 2016.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Wilkop T, Davis D, Pattathil S, Bao W, Peralta A, Domozych D, Hahn M, Drakakaki G. Endomembrane dynamics during cell division and plant morphogenesis 20th European Network for Plant Endomembrane Research meeting (ENPER). Prague, Czech Republic. September 12.- 15, 2017. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Davis D, Rosquete M, Wang H, Park E, Comai L, Domozych D, and Drakakaki G. Dissecting Cytokinesis in plants with ES7. Annual Meeting of the American Society of Plant Biologists Honolulu Hawai, USA. June 24-28, 2017. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Rosquete MR Worden N and Drakakaki G. ROG2, a novel player of Arabidopsis post-Golgi trafficking. UC Davis Postdoctoral Research Symposium. Davis, CA. April 2017. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Mabry E*, Rosquete M R, Drakakaki G. Cell Plate Formation and Mutant Plant Resistance to the Inhibitory Effects of ES7 Presented at the 28th Annual UC Davis Undergraduate Research Conference on April 28, 2017. *Undergraduate student presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Wang H-L, Davis D, Rosquete M, R�hrich R, Park E, Mackey S, McDowell S, Le T, Henry I, Comai L, and Drakakaki G. Using a small chemical to dissect callose deposition during cytokinesis, Annual Plant Biology Retreat, Marconi Conference Center, CA, USA. October 7-9 2016.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. The building up cell wall. Mediterean Agronomic Institute of Chania, Greece December 12, 2016
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Davis D. Dissecting cytokinesis in Plant Cells with ES7. Annual Meeting of the American Society of Plant Biologists Honolulu Hawai, USA June 24-28, 2017. Invited talk
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Endomembrane dynamics and genesis of cell wall. Shijiazhuang University, Hebei, China. July 21, 2017. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Endomembrane dynamics and polysaccharide deposition. The 6th International Conference on Plant Cell Wall Biology (PCWB2017) Dalian, China. July 16-20, 2017. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Towards a four dimensional model of cell plate formation. Workshop on quantitative cell biology: Finding your inner modeler. University of Illinois at Chicago, USA. July 13, 2017. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Davis DJ, Wang H. Rosquete MR, Park E, Wilkop T, Drakakaki G. Discovering how plant cells divide. NAIST International Student Workshop. November 16th 2016. Nara, Japan. Selected talk by D. Davis
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Dissecting post Golgi trafficking. Midwest Plant Cell Dynamics meeting Madison, USA. May 30th - June 2nd, 2017. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Drakakaki G. Probing endomembrane dynamics and polysaccharide deposition during the genesis of a new cell wall University of Minnesota, St Paul, MN, USA. April 25, 2017. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Davis DJ, Wang H, Wilkop T, Drakakaki, G. Plant cell cytokinesis with the small molecule inhibitor, Endosidin 7. UC Davis Plant Biology Retreat. Marconi Conference Center. October 7-9, 2016. Contributed talk by D. Davis.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Rosquete M.R, Worden N, Wilkop T, Drakakaki G. ROG2: Cell Plate Formation and Mutant Plant Resistance to the Inhibitory Effects of ES7. Annual Plant Biology Retreat, Marconi Conference Center, CA, USA. October 7-9, 2016. Contributed talk by Rosquete.


Progress 10/02/15 to 09/30/16

Outputs
Target Audience:The target audience for this project includes industry, commodity boards as well as scientists at UC Davis and off campus. The results were presented to a broader national and international audience in the forms of refereed publications, project reports, conference proceedings and oral presentations. Scientific community: Participants in the Gordon Research Conference on Plant & Microbial Cytoskeleton, XIV Cell Wall Meeting, NAIST International Student Workshop, Midwest Plant Cell Dynamics Meeting, scientists at Ohio University, the University of California Riverside and Kyushu University. The Almond and the Pistachio Research Boards. Scientists at UC Davis and participants at the Annual Almond Conference. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Within this project the following individuals were involved: Project scientist: Thomas Wilkop. Graduate students Natasha Worden and Destiny Davis. The postdoctoral researcher Dr. Michel Ruiz Rosquete , the undergraduate students: Shannon Meyhew, Nathan Jane, Emily Mabry, Sean Mackey, Marissa Huston, and the international visiting scholars: Hou-Ling Wang,Wenlong Bao, Yukun Cheng, Rachel Röhrich, Angelo Herringer, Alessandra Quartararo, Longjun Chen and the high-school student: John Almazan. How have the results been disseminated to communities of interest?We have disseminated our results in the form of publications, reports and scientific presentations. Examples of presentations to the scientific community include: Participants in the Gordon Research Conference on Plant & Microbial Cytoskeleton, XIV Cell Wall Meeting, NAIST International Student Workshop, Midwest Plant Cell Dynamics Meeting, scientists at Ohio University, the University of California Riverside and Kyushu University. The Almond and the Pistachio research boards. Scientists at UC Davis and participants at the Annual Almond Conference. What do you plan to do during the next reporting period to accomplish the goals?We will continue our efforts towards dissecting cell wall deposition and understanding plant response to biotic and abiotic stress.

Impacts
What was accomplished under these goals? The plant cell wall and its network of polysaccharides and glycoproteins is one of the evolutionarily conserved structures of terrestrial plants. Research in our laboratory is guided by the central hypothesis that transport and deposition of cell wall polysaccharides and biosynthetic enzymes occurs via specialized pathways within the endomembrane system. Given the complexity of cell wall synthesis and assembly, we are using a multidisciplinary approach including chemical genomics, classical genetics, proteomics and vesicle glycomics to establish in detail the mechanisms controlling trafficking and assembly of cell wall components and contribute to the build up of a new cell wall. 1) Plant cytokinesis, a process fundamentally different from cytokinesis in animals and fungi, requires the de novo formation of a cell plate that is maturing into a new cell wall and partitions the cytoplasm of the dividing cell. We identified and characterized Endosidin 7 (ES7) as a specific in vitro and in vivo inhibitor of callose deposition at the cell plate. The interplay between cell plate-specific post-Golgi vesicle traffic and callose accumulation under the influence of ES7 revealed unique and temporal contributions of secretory and endosomal vesicles in cell plate maturation. We identified and characterized mutants resistant to ES7, not exhibiting cell plate defects or a reduction of callose deposition at the cell plate upon ES7 exposure. Extending this genetic approach, we will identify new regulators of callose deposition and vesicle trafficking during cell plate formation. 2) Cell wall polysaccharides, associated enzymes, and glycoproteins must be transported through the secretory pathway to the specific sites of polysaccharide deposition into the cell wall. For a comprehensive understanding of polysaccharide deposition, detailed knowledge about the type of carbohydrate cargo delivered by transport vesicles is required. With the cell wall adapting to several biotic and abiotic stresses and stimuli, we investigate the endomembrane pathways contributing to these adaptations. We pioneered a method of vesicle isolation which we used to isolate post Golgi vesicles displaying the syntaxin of plants 61 (SYP61) and analyzed their proteome. Proteomic analysis suggested that SYP61 defines a pathway involved in cell wall deposition and plant stress response. Using our method of plant vesicle isolation in conjunction with a glycome profiling assay, we analyzed the polysaccharide content of SYP61 trans-Golgi vesicles. Our results provide for the first time a semi-quantitative, comprehensive list of plant polysaccharides transported in post Golgi vesicles. They demonstrate the degree of substitution in both xyloglucan and pectin existing in trans-Golgi vesicles compared to the cell wall. This provides insights into the type of modification that individual cell wall macromolecules undergo en route to the wall or during the process of cell wall assembly. 3) Pistachio and almonds are woody plants that are some of the major commercial crops in California. The current increase in global temperature in combination with intensive agriculture will substantially increase salt stress in crops. Soil salinization is a major challenge for most of the irrigated cropland in California. Understanding the mechanism of salt ion uptake, transport, and sequestration at the cellular and molecular levels can provide a way to identify desirable plant characteristics that can be selected for in rootstocks and scions in order to achieve optimal plant performance. Supported by the Almond and Pistachio Research Boards, we have developed a series of imaging tools and methodologies for the detection of sodium, potassium and chloride at the subcellular level with the aid of selective fluorescence indicator dyes that allow the quantitation of ion uptake and sequestration inside specific plant tissues. Our efforts contribute to a comprehensive characterization of complex mechanisms controlling salinity stress in woody plants.

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Davis D, Kang BH, Heringer AS, T.E. W, Drakakaki G (2016). Unconventional Protein Secretion in Plants. Methods Mol Biol. 2016;1459:47-63.
  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Bjornson M, Song X, Dandekar A, Franz A, Drakakaki G, Dehesh K. (2015). A Chemical Genetic Screening Procedure for Arabidopsis thaliana Seedlings. Bio Protoc. Jul 5;5(13). pii: e1519.
  • Type: Other Status: Published Year Published: 2016 Citation: Drakakaki G. 2016 Development of novel tools for dissection of salinity tolerance in almond. Cooperative personnel: Heringer A, Cheng Y, Aradhya M, Preece J, Lampinen B, Brown P, Gradziel T, Duncan R, Dorcey H, Wilkop T. Annual Report, Almond Research Board.
  • Type: Other Status: Published Year Published: 2016 Citation: Drakakaki G, Heringer A, Huston M, Ferguson L, Wilkop T. 2016. Cellular, subcellular and molecular characterization of salinity tolerance in pistachio with novel tools. Annual Report, Pistachio Research Board.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Wilkop T and Drakakaki G. Microscopy for the quantitative spatiotemporal analysis of cell plate growth. Workshop on the Quantitative Biology of Cytoskeletal Mechanics, Chicago, USA. October 22-24, 2015. Poster.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Drakakaki G. Dissecting post-Golgi trafficking during cytokinesis and cell wall formation. Plant Cell Biology retreat, Oct. 3, 2015. Davis, California
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. Post Golgi trafficking during cytokinesis and plant morphogenesis. Cell Biology Seminar Series, University of California Davis, Davis, California. April 8, 2016
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Jayne N*, Davis D, McDowell S, Park E, Drakakaki G Investigating Cell Plate Formation Using the Cytokinesis Inhibitor Endosidin 7. Presented at the 27th Annual UC Davis Undergraduate Research Conference, Davis CA. April 8, 2016. *Undergraduate student presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. Endomembrane dynamics and polysaccharide deposition during cell division and plant morphogenesis. Ohio University. April 18, 2016.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G Novel tool development in almond rootstocks. Almond Board rootstock research meeting. San Joaquin Valley Agricultural Sciences Center, Parlier California. April 26 2016 Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Davis DJ, Park E, McDowell S, Le T, Comai L, Drakakaki G. Using a small chemical to dissect polysaccharide deposition at the cell plate. Midwest Plant Cell Dynamics Meeting. St. Louis, Missouri. June 21-24, 2016. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Wilkop T, Davis D, Worden N, Pattathil S, Peralta A, Hahn M, Drakakaki G. Endomembrane dynamics and polysaccharide deposition during cell division and plant morphogenesis. XIV Cell Wall Meeting. Chania, Greece 12 - 17 June, 2016. Poster
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Davis DJ, Rosquete M.R, Wang H, Wilkop T, Drakakaki G. Dissecting plant cell cytokinesis with Endosidin 7. Midwest Plant Cell Dynamics Meeting, June 23, 2016. St. Louis, Missouri. Oral presentation by D. Davis.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Wilkop T and Drakakaki G. Dynamics of cell plate development during plant cytokinesis. 13th Annual Advanced Imaging Methods Workshop, UC Berkeley, California, USA. February 10 - 12, 2016. Poster.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. Cellular, subcellular and molecular characterization of salinity tolerance in pistachio with novel tools. Pistachio Research Board. Parlier CA, January 21, 2016 Oral presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Drakakaki G. Dissecting salinity tolerance in almonds with novel tools Annual Almond Conference, Sacramento, California, USA December 8 - 10, 2015. Oral presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Wilkop T, Heringer A, Le T, Esteva-Esteve V, Drakakaki G. Characterization of salinity tolerance in almonds with novel tools. Annual Almond Conference, Sacramento, California USA. December 8 - 10, 2015 Poster.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. Endomembrane dynamics during cytokinesis and cell elongation Plant & Microbial Cytoskeleton, Gordon research Conference, Proctor Academy, Andover, USA. August 14-19, 2016. Invited talk
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G Endomembrane trafficking during cytokinesis and cell elongation. University of California Riverside. March 20, 2016. Riverside CA. Invited talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Drakakaki G. Endomembrane dynamics during and cell division and plant morphogenesis Symposium on Organelle Homeostasis Research Center Protein Trafficking and Intracellular Signaling of Plant and Fungal Cells. Kyushu University School of Medicine, Fukuoka, Japan. February 8-9, 2016. Invited talk.