Progress 10/01/08 to 09/30/13
Outputs
Target Audience: The target audience are those interested in fundamental processes ongoing in seeds that influence seed dormancy and survival of, and longevity in, the dry state. We are examining the molecular level events important in both the protection of proteins from dehydration stress and the natural repair porcess that assist proteins in seeds recover from drastic water loss, recouping activity when subsequently rehydrated and repaired. These same processes also influence the dormancy exhibited by the seeds. Advances in understanding what specific proteins are protected from, or repaired after, dehydration stress will translate to disparate systems from pharmaceuticals to space flight.. The biotechnological benefit inherent in the capacity to remove water from functioning proteins, without catastrophic denaturation and aggregation, allowing the recapitulation of the functional form at some later time upon water addition is immense. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? The project dealt with protecting the seed proteome from dysfunction. Two of the post-docs hired on a similar project are now faculty (Dr. Tingsu Chen. Microbiology Institute, Guangxi Academy of Agricultural Sciences, Nanning, Guangxi, 530007, P. R. China; and Dr. Nihar Nayak. Regional Plant Resource Centre, Nayapalli, Bhubaneswar, Odisha, 751015, India). One high school student and one undergraduate researcher collaborated on work published in JBC 2010, a different undergraduate researcher collaborated on published work. How have the results been disseminated to communities of interest? Through six publications accepted in 2013. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Our discoveries of the preferred client proteins (CPs) for an orthologous pair of LEA proteins from the seed maturation protein (SMP) LEA family permitted mutant analysis of seeds deficient in each of the two most frequently acquired CPs. The seeds of mutants for either CP gene have the same heat-stress phenotype as when the LEA protecting them is dysfunctional. Modeling the CP sites to which these LEAs bind indicated a potential nucleic acid binding capacity, in keeping with gene ontology predictions and CP subcellular residence. This information, used in a suite of molecular and computational tools, provides insights into the LEA’s binding mechanism, the CPs’ functions, and the protein region bound by the LEA, a region that, by inference, may be the most susceptible portion of the CP to denaturation. The proposed studies will yield unprecedented insight as to how LEA proteins function in proteomic protection. The fundamental importance of maturation drying of the orthodox seed as the cornerstone of agriculture is in stark contrast to our lack of a basic understanding of the molecular mechanism of action for the LEA proteins, suspected for over 3 decades to be involved in the protection of the cellular milieu from stress. Through our efforts, these barriors in our knowledge are being removed. The Phage display libraries from this lab are being moved to the Arabidopsis Biological Resource Center, Ohio.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Kushwaha, Rekha, Payne, Christina, Downie, Bruce. 2013. Uses of phage display in Agriculture: A review of food-related protein-protein interactions discovered by biopanning over diverse baits. Computational and Mathematical Methods in Medicine. (Special Issue on Phage Display Informatics) Volume 2013, Article ID 653759, 12 pages. http://dx.doi.org/10.1155/2013/653759
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Nayak, Nihar, Putnam, Andrea, Chen, Tingsu, Sh�fermeyer, Kim R., Addepalli, Balasubrahmanyam, Lowenson, Jonathan, Jankowsky, Eckhard, Perry, Sharyn E., Dinkins, Randy, Limbach, Patrick, Clarke, Steven G., and Downie, A. Bruce. 2013. An ATP dependent DEAD-box RNA-helicase loses activity upon isoAsp formation but is restored by PROTEIN ISOASPARTYL METHYLTRANSFERASE. Plant Cell. 25:2573-2586.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Gu L, Han Z, Zhang L, Downie B, Zhao T. 2013. Functional analysis of the 5 regulatory region of the maize GALACTINOL SYNTHASE2 gene. Plant Science. 213: 38 45.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Petti C, Harman-Ware A E, Tateno M, Kushwaha R, Shearer A, Downie A B, Crocker M, DeBolt S. 2013. Sorghum mutant RG displays antithetic leaf shoot lignin accumulation resulting in improved stem saccharification properties. Biotechnology for Biofuels. 6: 146.
- Type:
Journal Articles
Status:
Accepted
Year Published:
2013
Citation:
Kushwaha, Rekha, Sch�fermeyer, Kim R., Downie, A. Bruce. 2013. A protocol for phage display and biopanning using recombinant protein baits. Journal of Visualized Experiments. in press.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Kushwaha, Rekha, Downie, Bruce, Payne, Christina. 2013. Uses of phage display in Agriculture: Sequence analysis and comparative modeling of Late Embryogenesis Abundant client proteins suggests protein-nucleic acid binding functionality. Computational and Mathematical Methods in Medicine. (Special Issue on Phage Display Informatics). Volume 2013, Article ID 470390, 12 pages. http://dx.doi.org/10.1155/2013/470390.
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: The PI spent a one year sabbatical in Dr. Frank Gubler's lab (host). Given a CSIRO protocol for germinating seeds. When adopted, using 0.7% w/v agarose and water only as media, a convincing inability of the ctg10 mutant to complete germination in the dark was consistently demonstrated over multiple generations and has persisted since my return to Kentucky. Similarly the RNAi lines are now behaving appropriately for their level of CTG10 reduction. Mentored Pratap Kumar Pati (PI) 2011-2012 during his Fulbright-Nehru Senior Research Fellowship. Polyadenylation of stored Messenger RNA during seed germination. August 2011-April 2012. PARTICIPANTS: Allan Bruce Downie and Enamul Huq lab (U Texas, Austin), have collaborated on an NSF grant. Frank Gubler: Hosted Downie for a 1 year sabbatical at CSIRO, Black Mountain, Canberra, Australia. Dr. Alejandra A. Covarrubias collaboration, Universidad Nacional Autonoma de Mexico, Mexico to use western blot (with antibodies to this LEA protein in her possession) to confirm that this LEA is persistent after imbibition, at least until the completion of germination. Santosh Kumar, Ph.D.: Dr. Kumar is the post-doc hired on PIF1 project. He is responsible for the generation and introduction into plants of the CTG10 RNAi constructs, and those N-TAPa tagged CTG10 constructs without an F-BOX. Rekha Kushwaha, Ph.D.: Dr. Kushwaha is a volunteer on this project. She is expressing recombinant PIF1 in E. coli. Upon purification, she has placed a defined amount of rPIF1 protein or BSA at the bottom of a microtiter plate well. She blocked the wells with milk powder or commercial blocking reagent and, after washing, added a given amount of the promoter of the gene encoding the DELLA protein, GA INSENSITIVE1 (GAI1), to which PIF1 is known to bind at the single G-Box in this promoter (pGAI). In other blocked wells containing BSA or PIF1, she introduced a mutated GAI promoter in which she had altered two bases in the G-box using site directed mutagenesis to a non-cannonical sequence. Mr. Kim Schafermeyer, B.Sc.: Kim assisted with the split YFP characterization of a PIF1/CTG10 interaction in vivo. Last year, Kim worked with Dr. Goodin and Ms. Martin to try to introduce the Agrobacterium carrying the CTG10 and PIF1 split YFP constructs into WT Nicotiana benthamiana leaves using infiltration and incubation. Kim also assisted in the use of biolistic introduction of the plasmids into tobacco leaf cells and confocal microscopy to follow the interaction of PIF1 with CTG10. Ms. Taylor Lloyd, undergraduate: One of the more exciting findings of Ms Lloyd's research was the identification of a significant alteration in CTG10 mRNA amounts in the pif1 KO and the PIF1 OE mutants relative to WT. CTG10 amounts in the pif1 KO were up relative to wild type while they were considerably repressed in the PIF1 OE line relative to WT. Examination of the promoter region of CTG10 showed that, while there were no G-boxes present, there were no fewer than 10 E-boxes. Ms Lloyd has recently been awarded the Astronaut Scholarship for the University of Kentucky. She continues to work on the CTG10-PIF1 project analysing PIF1 direct target gene expression using Q-RT-PCR under the auspices of a University of Kentucky Undergraduate Summer Research Grant. TARGET AUDIENCES: Research scientists. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
With Dr. Sharyn Perry, we collaborated with Mr. Brian Perry (Head naturalist at The Kentucky Nature Sanctuary, Raven Run) to host a laboratory and field program focusing on various ways scientists study and understand the biology and ecology of plant species. The original program (Mustards: From Mutations to Mutualism) is designed for late elementary school audiences and was successfully launched and presented four days between March 18th through the 25th in 2011. While the PI was on sabbatical (08.01.11-07.17.12), the program was run three days in 2012, and will be presented in 2013 for 3 days in the spring. The aim of this activity is to present science in a relaxed, field-day setting to economically underprivileged children to kindle within them the aspiration to pursue science as a career.
Publications
- Kushwaha, Rekha, Lloyd, Taylor, Schafermeyer, Kim R., Kumar, Santosh, Downie, A. Bruce. 2012. Identification of Late Embryogenesis Abundant (LEA) protein interactors using phage display. Int. J. Mol. Sci. 13, 6582-6603.
- Nosarzewski, Marta A C, Downie, A. Bruce, Wu, A. Benhong, Archbold, Douglas D. 2012. The role of SORBITOL DEHYDROGENASE in Arabidopsis thaliana. Functional Plant Biology. 39: 462-470.
- Taylor Lloyd et al..., Downie, AB. Two LEA homologues, retain a protein protective capacity in solution or when immobilized. ASPB 2012 Annual Meeting, Austin, Texas, July 20-24 2012.
- Rekha Kushwaha et al..., Downie, AB. Phage Display: An approach to identify Late Embryogenesis Abundant (LEA) interacting proteins. ASPB 2012 Annual Meeting, Austin, Texas, July 20-24 2012
- Bruce Downie et al. PROTEIN ISOASPARTYL METHYLTRANSFERASE rescues labile DEAD-box RNA helicase activity in vitro. ASPB 2012 Annual Meeting, Austin, Texas, July 20-24 2012
- Kill PIL. Arabidopsis COLD TEMPERATURE GERMINATING10 (CTG10) is a KELCH REPEAT CONTAINING F-BOX PROTEIN targeting PHYTOCHROME INTERACTING FACTOR 1 (PIF1 a.k.a. PIL5) and PIF3 for poly-ubiquitination and destruction. Dr. Anna Koltunow Hosting. CSIRO Plant Industry, Waite Campus, Adelaide, Australia June 26, 2012.
- Hormonal and Light Regulation of Grain Dormancy in Barley (Hordeum vulgare L.). CSIRO, Black Mountain Laboratories Hosting, June 28th, 2012
- RNA helicase activity is susceptible to isoAsp formation but recoverable by PROTEIN ISOASPARTYL METHYLTRANSFERASE. Bruce Downie, 16.02.2012, Plant Reproduction for Food 2012, Seed, Fruit and Seedless Fruit (C8), University of Melbourne, Melbourne, Australia, 13-17.02.2012
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: Objective 1. Identify and characterize biophysical, biochemical, genetic, and environmental factors regulation or influencing seed development, germination, vigor and dormancy. 1.a) Examination of the influence of barley (Hordeum vulgar) CRYPTOCHROME1 (CRY1), CRY2, LONG HYPOCOTYL5 (HY5), and COP INTERACTING PROTEIN8 (CIP8) on seed dormancy during germination. Objective 2. Determine and model the biotic and abiotic factors affecting seed germination, seedling emergence, and establishment of sustainable populations in natural and agro-ecological systems. 2.a) Used phage display and biopanning to examine the Arabidopsis LEA protein, SEED MATURATION PROTEIN1 (SMP1), and the soybean homolog, GmPM28, for protein binding partners at high temperature. 2.b) Used phage display and biopanning to examine PHYTOCHROME INTERACTING FACTOR1 (PIF1) interacting proteins in Arabidopsis thaliana. Recovery of interacting bHLH transcription factors to which PIF1 is known to bind as a homo- (PIF1) and a hetero- (PIF3) dimer and some to which PIF1 was not previously known to bind. The latter interactions were retrieved when genomic DNA containing E-BOX motifs was added to the pans. Objective 3. Develop, evaluate, and transfer technologies to assess and improve seed and seedling quality, health, performance, utilization, and preservation. No activity. Events include conferences, demonstration sites, field days, symposia, workshops, and trainings. Lloyd, Taylor, Kumar, Santosh, Downie, Bruce. Differential expression of PIF1-targeted genes in various PIF1 and CTG10 mutants. Abstract 517, ICAR, 2011, 22nd International Conference on Arabidopsis Research, June 22-25, Madison, Wisconsin, USA. Kushwaha, Rekha, Kumar, Santosh, Downie, Bruce. An approach to identify PHYTOCHROME INTERACTING FACTOR1 (PIF1) interacting proteins from Arabidopsis seeds. Abstract 512, ICAR, 2011, 22nd International Conference on Arabidopsis Research, June 22-25, Madison, Wisconsin, USA. Kumar, Santosh, Nayak, Nihar, Martin, Kathleen, Schafermeyer, Kim, Lloyd, Taylor D., Dinkins, Randy, Goodin, Michael, Downie, Bruce. Bimolecular Fluorescence Complementation studies support an in vivo interaction between the F-box protein COLD TEMPERATURE GERMINATING10 and PHYTOCHROME INTERACTING FACTOR1. Abstract 511, ICAR, 2011, 22nd International Conference on Arabidopsis Research, June 22-25, Madison, Wisconsin, USA. - Panel Member NSF-SBIR (2011: P110920. Phase I: Medical Imaging Technologies) - Associate Editor (seed biology) Botany. - Kentucky American - Mutant seed to Enamul, University of Texas, Austin, TX. - HsPIMT and PRH75 to Eckhard Jankowsky (Case Western Reserve University, Cleveland, Ohio. PARTICIPANTS: Participants: Funded project participants: Dr. Enamul Huq, University of Texas, is a co-PI on an NSF grant exploring the interaction of the PHYTOCHROME INTERACTING FACTOR1 (PIF1) and COLD TEMPERATURE GERMINATING10 (CTG10) proteins. This 2 year grant ended in 2011 but was granted a one-year no-cost extension until 2012. Dr. Arthur Hunt, University of Kentucky, is the PI on a joint project funded by the KSEF to explore the polyadenylation of transcripts in leaves and seeds of Arabidopsis. Dr. Eckhard Jankowski, Case Western Reserve University, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Andrea Putnam, Case Western Reserve University, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Balasubrahmanyam Addepalli, University of Cincinnati, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Jonathan Lowenson, University of California, Los Angeles, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Biranchi Patra, the KECK Institute, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Sharyn E. Perry, University of Kentucky, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Randy Dinkins, USDA-ARS Forage Animal Production Research Unit (FAPRU), is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Patrick A. Limbach, University of Cincinnati, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Animesh Ray, the KECK Institute, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Dr. Steven G. Clarke, University of California, Los Angeles, is a collaborator on a project exploring the PROTEIN ISOASPARTYL METHYLTRANSFERASE1 (PIMT1) target protein PLANT RNA HELICASE75 (PRH75). Late elementary school participants: Mustards: From Mutations to Mutualism: 2011 Raven Run Nature Sanctuary and College of Agriculture at the University of Kentucky collaborated on a new laboratory and field program focusing on various ways scientists study and understand the biology and ecology of certain species of plants. The program was presented for five days between March 18th through the 25th and was designed for late elementary school audiences. This program was sectioned into a laboratory component and a field component. Approximately 50 students attended each day of the program and were divided onto two groups. Each group began a different activity and switched after each hour to allow all students to complete both activities. TARGET AUDIENCES: The target audience for the research projects include all scientists with an interest in how seeds perceive the light environment (PIF1 CTG10 interaction) and the cellular response to hydration as it pertains to the polyadenylation or de-adenylation of stored or de novo synthesized transcripts. Concerning the project focused on the protection of the DEAD-box RNA helicase PRH75 by the PIMT1, this project has been completed and a paper written and submitted to a journal for publication. Regarding the outreach program of which I was a participant, the students of various middle schools were engaged in exploring the ecology of the plants of the mustard family at the Raven Run Nature Sanctuary and subsequently exposed to extraction and crude purification of genomic DNA from plant cells, a discussion of how plants perceive their environment, and a demonstration of how "mutant" plants may not be capable of correctly perceiving or interpreting, cues from their environment. This outreach program was undertaken with the view of increasing the awareness of middle school students of opportunities for careers in science. Mustards: From Mutations to Mutualism explored the many ways living organisms are studies by scientists from the genetic to the community level. Plants in the mustard family (Brassicaceae) were examined at the following levels: a. On the genetic level b. Organisms examined in a laboratory setting c. Populations on an ecological level d. Interactions between mustards and animal species at a community level e. Human relationships to mustard plants The main goals of this program was to provide students with: a. Hands-on experiences of the methods scientists use to study organisms b. Presentations answering the following questions a. How do scientists study questions related to plant genetics and adaptations in a laboratory using the scientific method b. What is DNA and how is the genetic information of mustards related to adaptations that we can observe in these plants c. What types of mustard plants can be found in forest and meadow ecosystems in Kentucky and how do they use the physical conditions of their environment, such as sunlight, to determine when to flower d. How do the relationships between mustards and certain animals illustrate the adaptations found in each species e. What type of mustard plants do people use and how are they related to the mustards found in Kentucky forests and meadows c. A field experience that allowed students to explore the forests and meadows of Raven Run Nature Sanctuary d. Access to professional scientists and naturalists who addressed student's questions concerning biology, methods of sciences and career activities. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
As a direct result in participating in research activities in my lab, Ms. Taylor Lloyd's scientific capacity increased to the point where she was capable of acquiring her own funding in the form of an American Society of Plant Biologists Summer Undergraduate Research Fellowship and the Astronaut Fellowship. Former post-doc Dr. Nihar Ranjan Nayak got a job as a Ramalingaswami Fellow at the Regional Plant Resource Center, Nayapalli, Bhubaneswar, Odisha,751 015, India. Former post-doc Dr. Tianyong Zhao is now Professor in the Department of Biochemistry and Molecular Biology, College of Life Science, Northwest A&F University, Yangling, Shannxi 712100, P.R.China. Seeking to perform a Sabbatical in Australia starting in August of 2011, the following funds were acquired: Office of the Chief Executive Science team: Commonwealth Science and Industry Research Organization (CSIRO): Sir Frederick McMaster Fellowship. 01/08/2011-31/07/2012 ($15,300 AUD). USDA-Strengthening Grant: Sabbatical: 2011-04375. Using RNA interference (RNAi) to deconvolute cryptochromes' (HvCRYs) influences on gene expression controlling ABA titer impacting seed dormancy in Barley (Hordeum vulgare). 01.02.2012-31.01.2013. ($64,934 for one year) From a collaboration with Arthur Hunt of Plant and Soil Science, the funds for a second year of a two year grant were provided for the continuence of a KSEF grant. Kentucky Science and Engineering Foundation (KSEF through KSTC) ($100,000). PI: Arthur Hunt. Co-PI: Bruce Downie. Polyadenylation of stored mRNA during seed germination. July 01, 2010-June 30, 2012.
Publications
- Mendu Venugopal, Griffiths Jonathan S., Persson Staffan, Stork Jozsef, Downie A. Bruce, Voiniciuc Cătălin, Haughn George W., DeBolt Seth. 2011. Subfunctionalization of cellulose synthases in seed coat epidermal cells mediates secondary radial wall synthesis and mucilage attachment. Plant Physiol. 157: 441-453.
- Xiaohui Wu, Qingshun Li, Bruce Downie, Chun Liang, Guoli Ji, and Arthur Hunt. 2011. Genome-wide landscape of polyadenylation in Arabidopsis provides evidence for extensive alternative polyadenylation. PNAS USA. 108: 12533-12538.
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: Our overall goal is to understand the ecology, biogeography and evolution of seed dormancy and germination. A concentrated effort to examine the seed dormancy mechanism of species present in biogeographical regions identified as underrepresented in the 1998 book Seeds: Ecology, biogeography, and evolution of dormancy and germination. has resulted in publications on several species from the USA as well as species from other countries including Taiwan, Japan, China, Australia (Dodonaea), and Europe (forest herbs). Web page for the W-2168 was set up that links to all member's labs (http://www.seedresearchers.com). The Perry lab has continued to probe mechanisms underpinning plant embryogenesis by identifying genes directly and indirectly regulated by the DNA-binding MADS-factor AGL15. AGL15 accumulation correlates with competence for somatic embryogenesis. Understanding somatic embryo development is fascinating from the basic viewpoint of determining how a cell can dedifferentiate and redifferentiate in embryo mode, as well as having practical aspects to promote regeneration of value-added transgenic plants by somatic embryogenesis. One route to contribute to determination of mechanisms of somatic embryo development is to determine genes controlled by transcriptional regulators that promote this process, such as AGL15. We have globally mapped where AGL15 associates with DNA in vivo, resulting in ~2000 binding sites. We have combined this information with expression microarray data to determine perturbations in the transcriptome in response to increased or decreased AGL15. This has allowed us to identify genes that are likely to be directly expressed in response to AGL15 as well as genes directly repressed by AGL15, resulting in ~200 genes with a direct, consistent and significant response to AGL15. Interestingly, other transcriptional regulators are overrepresented in the directly expressed list, but not the directly repressed list. DNA motifs that AGL15 recognizes are overrepresented in the fragments identified as bound by AGL15. Currently we are focusing on confirmation of select direct targets of AGL15 (and the redundant AGL18) and to determine the function of regulated genes. Finally we are developing tools to allow us to extend the gene regulation network underlying embryogenesis by looking at genes regulated by some AGL15 targets that encode transcriptional regulators. The data obtained has also led to an ongoing collaboration with a researcher at the University of Wisconsin-Madison. PARTICIPANTS: An Agriculture Biotechnology Student (Taylor Lloyd) and a high school Student (Alyssa Eliopolous), Enamul Huq (University of Texas as Austin), fullbright fellowship for a visiting scientist from India to have an 8 month sojourn in my lab working on this topic, a new graduate student in the Hunt lab (Liuyin Ma) is dedicated to this project as his Ph.D. topic. This is funded by the Kentucky Science and Engineering Foundation. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
A full-day workshop entitled "A short-course on seed dormancy and germination with emphasis on Hawaiian species" University of Hawaii, Manoa. Half-day workshop entitled "Future research objectives for studies on seed dormancy and germination in the Junggar Desert, northwest China." Xinjiang Agriculture University, Urumqi, China. Lecture: "Seed dormancy profile for the Xishuangbanna Tropical Seasonal Rainforest Dynamics Plot: First approximation" Chinese Academy of Sciences, Xishuangbanna Tropical Botanical Garden, Kunming, China. Lecture. "Variation in seed dormancy/germination within and between individuals and populations" Chinese Academy of Sciences, Xishuangbanna Tropical Botanical Garden, Kunming, China. Lecture. " The ecology of the federally-endangered species Solidago shortii" Xinjiang Agricultural University, Urumqi, China. Lecture. "How seed dormancy studies serve plant conservation: Using information on biogeography and phylogeny of seed dormancy to facilitate propagation of plant species for restoration" Lyon Arboretum, University of Hawaii, Manoa. Symposium talk. "Seed dormancy and germination of Hawaiian montane species: meeting common goals of basic science and conservation" Hawaii Conservation Conference, Honolulu, HI. Lecture. "Seed germination ecology in a specific habitat: Cedar glades" Xinjiang Agricultural University, Urumqi, China. Lecture. "Seed dormancy and germination of cedar glade plants" Cedars of Lebanon Annual Wildflower Weekend, Lebanon, TN. Invited poster. "Classification and phylogeny of seed dormancy", Xishuangbanna International Symposium II. Biodiversity Conservation: Research Imperatives for Scientific Institutions" Chinese Academy of Sciences, Institute of Botany, Xishuangbanna Tropical Botanical Garden, Menglum (Yunnan Province), P. R. China. The membership was expanded, an official name for the W-2168, that will not change each time we go through a re-write, was adopted, a webpage for the group was established. We have now published our results adding to an emerging picture of direct transcriptional control. We have found, as have others, that while many sites are occupied by a given transcription factor in vivo (generally 1000s of DNA binding sites), relatively few interactions between a transcriptional regulator and DNA result in significant changes in gene expression of the nearby gene. Analysis of genes directly and indirectly controlled by AGL15 has led us to investigate hormone interactions involved in control of somatic embryo development. We are currently using combinations of mutants, hormones and inhibitor treatments to tease apart how different hormoneal cross-talk controls somatic embryo development. We have extended our work in Arabidopsis and demonstrated that ectopic expression of AGL15 in soybean can promote somatic embryogenesis in this important crop.
Publications
- Chen, Tingsu, Nayak, Nihar, Maitra Majee, Susmita, Lowenson, Jonathan, Schafermeyer, Kim R., Eliopoulos, Alyssa C., Lloyd, Taylor D., Dinkins, Randy, Perry, Sharyn E., Forsthoefel, Nancy R., Clarke, Steven G., Vernon, Daniel M., Zhou, Zhaohui Sunny, Rejtar, Tomas, and Downie, A. Bruce. 2010. Substrates of the Arabidopsis thaliana PROTEIN ISOASPARTYL METHYLTRANSFERASE 1 identified using phage display and biopanning. Journal of Biological Chemistry. 285: 37281 to 37292.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: Our overall goal is to understand the ecology, biogeography and evolution of seed dormancy and germination. A concentrated effort to examine the seed dormancy mechanism of species present in biogeographical regions identified as underrepresented in the 1998 book, Seeds: Ecology, biogeography, and evolution of dormancy and germination has resulted in publications in 2009 on several species from the USA as well as species from other countries including Taiwan, Japan, China, Australia Dodonaea), and Europe (forest herbs). Web page for the W-2168 was set up that links to all member's labs (http://www.seedresearchers.com). The Perry lab has continued to probe mechanisms underpinning plant embryogenesis by identifying genes directly and indirectly regulated by the DNA-binding MADS-factor AGL15. AGL15 accumulation correlates with competence for somatic embryogenesis. Understanding somatic embryo development is fascinating from the basic viewpoint of determining how a cell can dedifferentiate and redifferentiate in embryo mode, as well as having practical aspects to promote regeneration of value-added transgenic plants by somatic embryogenesis. One route to contribute to determination of mechanisms of somatic embryo development is to determine genes controlled by transcriptional regulators that promote this process, such as AGL15. We have globally mapped where AGL15 associates with DNA in vivo, resulting in around 2000 binding sites. We have combined this information with expression microarray data to determine perturbations in the transcriptome in response to increased or decreased AGL15. This has allowed us to identify genes that are likely to be directly expressed in response to AGL15 as well as genes directly repressed by AGL15, resulting in around 200 genes with a direct, consistent and significant response to AGL15. Interestingly, other transcriptional regulators are overrepresented in the directly expressed list, but not the directly repressed list. DNA motifs that AGL15 recognizes are overrepresented in the fragments identified as bound by AGL15. We published this work in The Plant Cell in 2009. Currently we are focusing on confirmation of select direct targets of AGL15 (and the redundant AGL18) and to determine the function of regulated genes. Finally we are developing tools to allow us to extend the gene regulation network underlying embryogenesis by looking at genes regulated by some AGL15 targets that encode transcriptional regulators. The data obtained has also led to collaborations resulting in a publication in the Journal of Experimental Botany in 2009 and an ongoing collaboration with a researcher at the University of Wisconsin-Madison. PARTICIPANTS: PI: Carol Baskin, PI: Jerry Baskin, PI: Lynnette Dirk, PI: Bruce Downie, PI: Robert Geneve, PI: Sharyn Perry, Collaborator: C. Barton, Collaborator: Z. Cao, Collaborator: O. Chabrerie, Collaborator: S. Y. Chen, Collaborator: W. Chen Collaborator: C. T. Chien, Collaborator: L. E. Commander, Collaborator: A. Cook, Collaborator: G. Decocq, Collaborator: M. Diekmann, Collaborator: K. W. Dixon, Collaborator: M. Dong, Collaborator: G. Z. Du. Collaborator: B. J. Graae, Collaborator: T. Heinken, Collaborator: M. Hermy, Collaborator: Y. Hoshino, Collaborator: X. W. Hu, Collaborator: Z. Huang, Collaborator: D.T. Karlson, Collaborator: E. Karlsson, Collaborator: A. Kolb, Collaborator: T. Kondo, Collaborator: J.A. Long, Collaborator: J. J. Lu, Collaborator: K. Nakaminami, Collaborator: S. S. Phartyal, Collaborator: N. Ren, Collaborator: N. Sentoku, Collaborator: K. J. Steadman, Collaborator: L. Strom, Collaborator: A.J. Stromberg, Collaborator: H. Z. Sun, Collaborator: D. Y. Tan, Collaborator: Y. Tang, Collaborator: D. M. TeKrony, Collaborator: S. R. Turner, Collaborator: K. Valtinat, Collaborator: S. Van Der Veken, Collaborator: K. Verheyen, Collaborator: H. Wang, Collaborator: J. H. Wang, Collaborator: Y. R. Wang, Collaborator: Y. P. Wu Collaborator: G. Yang, Collaborator: L. Yang. Collaborator: S. Zhang, Collaborator: X. Zhu, Collaborator: Z. Zheng, Collaborator: R. Zindel, Student: K. Hill, Student: K. M. G. G. Jayasuriya, Student: H. Yang, Student: Y. Zheng, TARGET AUDIENCES: Seed biologists worldwide. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Workshop "A short-course on seed dormancy and germination with emphasis on Hawaiian species" University of Hawaii, Manoa, 31 July 2009. Workshop "Future research objectives for studies on seed dormancy and germination in the Junggar Desert, northwest China." Xinjiang Agriculture University, Urumqi, China, 27 June 2009. Lecture: "Seed dormancy profile for the Xishuangbanna Tropical Seasonal Rainforest Dynamics Plot: First approximation" Chinese Academy of Sciences, Xishuangbanna Tropical Botanical Garden, Kunming, China. 24 December 2009. Lecture. "Variation in seed dormancy/germination within and between individuals and populations" Chinese Academy of Sciences, Xishuangbanna Tropical Botanical Garden, Kunming, China. 24 December 2009. Lecture. "The ecology of the federally-endangered species Solidago shortii" Xinjiang Agricultural University, Urumqi, China. 17 June 2009. Lecture. "How seed dormancy studies serve plant conservation: Using information on biogeography and phylogeny of seed dormancy to facilitate propagation of plant species for restoration" Lyon Arboretum, University of Hawaii, Manoa, 3 August 2009. Symposium talk. "Seed dormancy and germination of Hawaiian montane species: meeting common goals of basic science and conservation" Hawaii Conservation Conference, Honolulu, HI, 30 July 2009. Lecture. "Seed germination ecology in a specific habitat: Cedar glades" Xinjiang Agricultural University, Urumqi, China. 17 June 2009. Lecture. "Seed dormancy and germination of cedar glade plants" Cedars of Lebanon Annual Wildflower Weekend, Lebanon, TN, 2 May 2009. Invited poster. "Classification and phylogeny of seed dormancy", Xishuangbanna International Symposium II. Biodiversity Conservation: Research Imperatives for Scientific Institutions" Chinese Academy of Sciences, Institute of Botany, Xishuangbanna Tropical Botanical Garden, Menglum (Yunnan Province), P. R. China, 1-2 January 2009. The membership was expanded, an official name for the W-2168 that will not change each time we go through a re-write, was adopted, a webpage for the group was established. Limited numbers of studies have been performed to globally identify direct targets of plant transcription factors. Consequently not much is known about the number or types of targets that a specific factor controls. We have found that while many sites are occupied by a given transcription factor in vivo relatively few interactions between a transcriptional regulator and DNA result in significant changes in gene expression of the nearby gene. There are numerous possible reasons, but it seems important to assess the impact of DNA-protein interaction. Analysis of genes directly and indirectly controlled by AGL15 has led us to investigate hormone interactions involved in control of somatic embryo development. We are currently using combinations of mutants, hormones and inhibitor treatments to tease apart how different hormones cross-talk in control of somatic embryo development. We have extended our work in Arabidopsis and demonstrated that ectopic expression of AGL15 in soybean can promote somatic embryogenesis in this important crop.
Publications
- Baskin, C. C., C. T. Chien, S. Y. Chen and J. M. Baskin. 2009. Epicotyl morphophysiological dormancy in seeds of Daphniphyllum glaucescens, a woody member of the Saxifragales. International Journal of Plant Sciences 170: 174-181.
- Jayasuriya, K. M. G. G., J. M. Baskin, R. L. Geneve, C. C. Baskin. 2009. Phylogeny of Seed Dormancy in Convolvulaceae, Subfamily Convolvuloideae (Solanales). Annals of Botany 103: 45-63.
- Jayasuriya, K. M. G. G., J. M. Baskin, R. L. Geneve, C. C. Baskin. 2009. A proposed mechanism for physical dormancy break in seeds of Ipomoea lacunosa (Convolvulaceae). Annals of Botany 103: 433-445.
- Phartyal, S. S., T. Kondo,Y. Hoshino, C.C. Baskin and J.M. Baskin. 2009. Morphological dormancy in seeds of the autumn-germinating shrub Lonicera caerulea var. emphyllocalyx (Caprifoliaceae). Plant Species Biology 24: 20-26.
- Jayasuriya, K. M. G. G., J. M. Baskin, R. L. Geneve and C. C. Baskin. 2009. Sensitivity cycling and mechanism of physical dormancy break in seeds of Ipomoea hederacea (Convolvulaceae). International Journal of Plant Sciences 170: 429-443.
- Jayasuriya, K. M. G. G., J. M. Baskin and C. C. Baskin. 2009. Sensitivity cycling and its ecological role in seeds with physical dormancy. Seed Science Research 19: 3-13.
- Hu, X.W., Y. R. Wang, Y. P. Wu and C.C. Baskin. 2009. Role of the lens in controlling water uptake in seeds of two Fabaceae (Papilionoideae) species treated with sulphuric acid and hot water. Seed Science Research 19: 73-80.
- Jayasuriya, K. M. G. G., J. M. Baskin, D. M. TeKrony, and C. C. Baskin. 2009. Sensitivity cycling to physical dormancy break and seed vigour of two Ipomoea species (Convolvulaceae). Seed Science Research 19: 249-259.
- Hall, S., C. Barton, C. C. Baskin. 2009. Topsoil seed bank of an oak-hickory forest in eastern Kentucky as a Restoration Tool on Surface Mines. Restoration Ecology DOI: 10.1111/j.1526-100X.2008.00509.x
- Hall, S. L., C. D. Barton and C. C. Baskin. 2009. Seed viability in stockpiled topsoil on a surface mine in Appalachia. Ecological Restoration 27:381-383.
- Wang, J. H., C. C. Baskin, W. Chen and G. Z. Du. 2009 Variation in seed germination between populations of five sub-alpine woody species from eastern Qinghai-Tibet Plateau following dry storage at low temperatures. Ecological Research DOI 10.1007/s11284- 009-0643-0
- Zheng, Z., S. Zhang, G. Yang, Y. Tang, J. M. Baskin, C. C. Baskin and L. Yang. 2009. Abundance and distribution of cavity trees in an old-growth subtropical montane evergreen broad-leaved forest. Canadian Journal of Forest Research 39: 2234-2245.
- Graae, B. J., K. Verheyen, A. kolb, S. Van Der Veken, H. Heinken, O. Chabrerie, M. Diekmann, K. Valtinat, R. Zindel, E. Karlsson, L. Strom, G. Decocq, M. Hermy and C. C. Baskin. 2009. Germination requirements and seed mass of slow- and fast-colonizing temperate forest herbs along a latitudinal gradient. Ecoscience 16: 248-257.
- Chen, S.-Y., Chien, C.-T., Baskin, J. M. and Baskin, C. C. 2009. Storage behavior and changes in concentrations of abscisic acid and gibberellins during dormancy break and germination in seeds of Phellodendron amurense var. wilsonii (Rutaceae). Tree Physiology 30: 275-284.
- Geneve, R.L. (2009). Physical seed dormancy in selected Caesalpinioid legumes from eastern North America. Propagation of Ornamental Plants 9:129-134.
- Jayasuriya, K.M.G., Baskin, J.M., Geneve, R.L. and Baskin, C.C. (2009). A proposed mechanism of physical dormancy break in sensitive and insensitive seeds of Ipomoea lacunosa (Convolvulaceae). Annals of Botany 103:433-445.
- Jayasuriya, K.M.G., Baskin, J.M., Geneve, R.L. and Baskin, C.C. (2009). Phylogeny of seed dormancy in Convolvulaceae, subfamily Convolvuloideae (Solanales). Annals of Botany 103:45-63.
- Yang, H., C.C. Baskin, J. M. Baskin, Z. Cao, X. Zhu1, Z. Huang, M. Dong. 2009. Responses of caryopsis germination, early seedling growth and ramet clonal growth of Bromus inermis to soil salinity. Plant and Soil 316: 265-275.
- Phartyal, S. S. T. Kondo, J. M. Baskin and C.C. Baskin. 2009. Temperature requirements differ for the two stages of seed dormancy-break in Aegopodium podagraria (Apiaceae), A species with deep complex morphophysiological dormancy. American Journal of Botany 96: 1086-1095.
- Graae, B. J., K. Verheyen, A. Kolb, S. Van Der Veken, T. Heinken, O. Chabrerie, M. Diekmann, K. Valtinat, R. Zindel, E. Karlsson, L. Strom, G. Decocq, M. Hermy and C.C. Baskin. 2009. Germination requirements and seed mass of slow- and fast-colonizing temperate forest herbs along a latitudinal gradient. Ecoscience 16: 248-257.
- Baskin, C. C., S.-Y. Chen, C.-T. Chien and J. M. Baskin. 2009. Overview of seed dormancy in Viburnum (Caprifoliaceae). Propagation of Ornamental Plants 9: 115-121.
- Sun, H. Z., J. J. Lu, D. Y. Tan, J. M. Baskin and C. C. Baskin. 2009. Dormancy and germination characteristics of the trimorphic achenes of Garhadiolus papposus (Asteraceae), an annual ephemeral from the Junggar Desert, northwest China. South African Journal of Botany 75: 537-545.
- Turner, S. R., A. Cook, J. M. Baskin, C. C. Baskin, K. J. Steadman and K. W. Dixon. 2009. Identification and characterization of the water gap in the physically dormant seeds of Dodonaea petiolaris: a first report for Sapindaceae. Annals of Botany 104: 833-844.
- Turner, S. R., L. E. Commander, J. M. Baskin, C. C. Baskin and K. W. Dixon. 2009. Germination behaviour of Astroloma xerophyllum (Ericaceae), a species with woody indehiscent endocarps. Botanical Journal of the Linnean Society 160: 299-311.
- Zheng, Y., Ren, N., Wang, H., Stromberg, A.J. and Perry, S.E. 2009. Global Identification of Targets of the Arabidopsis MADS Domain Protein AGAMOUS-Like15. The Plant Cell 21, 2563-2577.
- Nakaminami, K., Hill, K., Perry, S.E., Sentoku, N., Long, J.A. and Karlson, D.T. 2009. Arabidopsis Cold Shock Domain Proteins: Relationships to Floral and Silique Development. Journal of Experimental Botany 60, 1047-1062.
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