Progress 03/01/10 to 09/30/14
Outputs
Target Audience: The target audience includes plant and animal biologists. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? This project provided opportunities for postdoctoral, graduate and undergraduate students training in laboratory research and public presentations. In addition, this project provided a training opportunity to an international intern student. How have the results been disseminated to communities of interest?
Nothing Reported
What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
We characterized the roles of the FT/TFL1 gene family in photoperiodic flowering in Arabidopsis. Using an RNA-sequencing approach, we analyzed mRNA expression data to identify genes potentially regulated by TFL1. Known targets of TFL1 as well as potentially further downstream targets were found. Using a transgenic approach based on two induction systems, we aimed to identify the immediate targets of FT and TFL1. The results of this approach suggest that FT controls downstream genes in an indirect manner. In addition, we have characterized the functions of soybean FT homologs, particularly in relation to soybean development and evolution. We focused on one of the FT homologs and found that mRNA expression data, gene structure data and functional data suggest that this gene stimulates flowering in wild soybean but is nonfunctional in domesticated soybean.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Wang, M. and Hanzawa, Y. Comparative transcriptome analysis of photoperiodic flowering response of maturity loci (E Loci) in soybean. Molecular and Cellular Biology of Soybean 2014 (Minneapolis, MN).
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Wu, F., Price, B., Haider, W., Seufferheld, G., Nelson, R. and Hanzawa, Y. Functional and evolutionary characterization of the CONSTANS gene family in short-day photoperiodic flowering in soybean. Molecular and Cellular Biology of Soybean 2014 (Minneapolis, MN).
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Sedivy, E. and Hanzawa, Y. Association mapping of temperature response in wild soybean Glycine soja. Molecular and Cellular Biology of Soybean 2014 (Minneapolis, MN).
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Rosas, U., Mei, Y., Xie, Q., Banta, J., Seufferheld, G., Gerald-Martinez, S., Chou, L., Bhambra, N., Parks, J.D., Flowers,
J., McClung, C.R., Hanzawa, Y. and Purugganan, M.D. Variation in Arabidopsis flowering time associated with cis-regulatory variation in CONSTANS. Nature Commun. 5:3651-3658.
- Type:
Journal Articles
Status:
Accepted
Year Published:
2015
Citation:
Wickland, D.P. and Hanzawa, Y. The flowering locus T/Terminal Flower 1 gene family: Functional evolution and molecular mechanisms. Molecular Plant.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2015
Citation:
Sedivy, E.J., Mei, Y., Donahue, J., Nakamura, Y., Teo, N.Z.W., Wenk, M.R., Ito, T., Bradley, D., Gillaspy, G. and Hanzawa, Y. Phospholipid signaling modulates flowering and inflorescence architecture controlled by TFL1 in Arabidopsis.
- Type:
Theses/Dissertations
Status:
Published
Year Published:
2014
Citation:
Wickland, D.P. Functional characterization of the functional characterization of the FT/TFL1 gene family in photoperiodic flowering in Arabidopsis and soybean.
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Progress 01/01/13 to 09/30/13
Outputs
Target Audience: The target audience includes plant and animal biologists. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Our progress obtained in 2013 provided indirectly training opportunities for one postdoctoral researcher, 3 graduate students and 2 undergraduate students. One of these graduate students defended a M.S. and currently is a Ph.D. student in my lab, and another student defended a Ph.D. The graduate students attended a scientific conference and gave three poster presentations. How have the results been disseminated to communities of interest? The results obtained through this project have produced a research publication in a peer reviewed journal, as well as a book chapter in a text book. In addition, the results and achievements obtained through this project produced a basis of three public poster presentations in a scientific conference. Moreover, the PI had opportunities to present the achievement through this project in two invited lectures. What do you plan to do during the next reporting period to accomplish the goals? I plan to develop more of the application side of this project, as well as produce more publications out of this project.
Impacts
What was accomplished under these goals?
TFL1 and FT are key flowering regulators that control the activity of the shoot apical meristem (SAM) in flowering plants. We previously identified TIL3, encoding a phosphatase, as a TFL1-interacting protein in vitro. During 2013, we demonstrated the role of TIL3 in TFL1-regulated flowering control. We confirmed the TFL1-TIL3 protein-protein interaction in vivo with transiently expressed bimolecular complementation (BiFC) in Nicotiana benthamiana. Both 35S:GFP:TFL1 and 35S:GFP:TIL3 showed localization in the nucleus and the endoplasmic reticulum. In addition, til3 loss-of-function mutants and its homologs til3like1 and til3like2 were observed to reduce the late-flowering phenotype of 35S:TFL1 transgenic plants. Furthermore, both 35S:TFL1 and mutant tfl1-1 display increased sensitivity to gravity in root gravitropism similar to the til3 mutant. We conclude that 5Ptase13 modulates floral regulation through direct interactions with TFL1. Aiming at clarification of TFL1/FT gene family in soybean, we cloned full-length cDNAs of 3 soybean FT homologs. These cDNAs are currently being transformed into Arabidopsis to examine their function in flowering control.
Publications
- Type:
Journal Articles
Status:
Accepted
Year Published:
2013
Citation:
Wu, F., Price, B.W., Haider, W., Seufferheld, G., Nelson, R. and Hanzawa, Y. Functional and evolutionary characterization of the CONSTANS gene family in short-day photoperiodic flowering in soybean. PLOS ONE.
- Type:
Book Chapters
Status:
Awaiting Publication
Year Published:
2014
Citation:
Wu, F. and Hanzawa, Y. Photoperiodic control of flowering in plants. In: Pessarakli M, editor. Handbook of plant and crop physiology. 3rd ed. Boca Raton, Florida: CRC press. pp. 129-139.
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: To clarify the molecular actions of key flowering genes FT and TFL1, we have characterized the function of TIL3, a TFL1 interacting protein, and its homologs. Aiming at confirming TFL1 and TIL3 interactions in plants, we produced transgenic Arabidopsis plants that express chimeric proteins: TFL1:GFP, GFP:TFL1, TIL3:RFP, and RFP:TIL3. In addition, we have taken a tobacco transient expression system to express these proteins. Our observation on conforcal microscopy indicated that TFL1 and TIL3 co-localize in the same cellular organelle: overlapping signals were detected in nucleus and cytosol. This result indicates TFL1 and TIL3 may indeed interact in plants. To further clarify their interaction, we are currently carrying out a co-IP experiment. Furthermore, to visualize the interaction, we are subcloning TFL1 and TIL3 in BiFC vectors that allow detection of protein-protein interactions by emitting fluorescent signals to observe under microscopy. Our second approach to FT and TFL1 is clarification of downstream genes. Previously, we identified a set of genes that is up-regulated by FT and down-reguated in TFL1, indicating the antagonistic actions of FT and TFL1 in downstream gene regulation. To test whether the set of genes is immediate downstream of TFL1, we are currently developing inducible TFL1 Arabidopsis transgenic plants. Moreover, our collaboration with Prof. Huber (USDA) indicated plant hormone pathways and the TFL1 pathway may overlap. We have initiated further investigation on this aspect using molecular genetics and biochemical approaches. Our ultimate goal is to transfer the knowledge obtained in a model species to economically important crop plants. To achieve this goal, we comparatively work on soybean, the second most produced crop in the U.S. We have collected shoot materials from soybean plants that were treated with different day-length conditions and carried out an expression analysis of key flowering genes using quantitative RT-PCR and transcriptome sequencing. G. max reference varieties, near-isogenic lines of E loci (E1, E2, E3, and E5), and a soybean ancestor, G. soja are used for this study. E loci responded differently to day-lengths and time points. GO term analysis showed enrichment of different functions in different E loci. Key flowering gene homologs in the photoperiod pathway, including GmCO and GmFT, showed a different expression pattern from that of Arabidopsis CO and FT, whereas genes in the circadian clocks and the vernalization pathways showed a similar expression pattern. The peak expressions of GmCO2a and GmCO2b overlap with that of GmFT, suggesting that they might be a regulator of GmFT. Regulatory interactions of flowering genes are currently being elucidated further using a co-expression-based network analysis. Our additional approach to soybean flowering mechanisms is to identify novel genes underlying photoperiodic flowering in soybean by exploring the G. soja genome. Our QTL mapping analysis identified previously unidentified flowering and maturation loci. PARTICIPANTS: Potential collaborators: Randall Nelson, USDA and Department of Crop Sciences, University of Illinois. Matthew Hudson, Department of Crop Sciences, University of Illinois. Steven Huber, USDA, University of Illinois. Michael D. Purugganan, Department of Biology, New York University. Desmond Bradley, John Innes Centre, UK. Francisco Madueno, University of Valencia, Spain. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Our results and achievements obtained in 2012 became a basis of five public poster presentations in scientific conferences and two invited seminars domestic and abroad. These presentations significantly enhanced the recognition of the scholarly activities of the PI and the University in public. These activities provided the PI opportunities to develop new collaborative interactions with researchers in other institutions. In addition, we are currently preparing two manuscripts to submit Dec 2012 - January 2013, and further 2 research papers to submit in early 2013.
Publications
- No publications reported this period
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: To understand the function of key flowering genes and their interactions, multiple strategies are being taken. Summary outputs during the period of 1/1/2011 - 12/31/2011 are described below. Homologous proteins FT and TFL1 are essential for flowering control in plants, but their actions are antagonistic. To clarify the molecular actions of the FT and TFL1, we isolated a TFL1 interacting protein TIL3. We have isolated a T-DNA insertion mutant of TIL3 and that of its homologs, TIL3L1, TIL3L2, and TIL3L3 to examine their function in flowering control. Although the single mutants did not show any statistically significant effects on flowering time, til3, til3l1, and til3l2 mutants turned out to repress the late flowering effect of the TFL1-over expressing plants 35S::TFL1, suggesting that the TIL3 gene family acts in the downstream of TFL1. To test the interaction of TFL1 and the TIL3 family proteins, we have created transgenic Arabidopsis plants expressing TFL1 tagged with GFP or the TIL3 family protein tagged with RFP, and currently crossing these plants aiming for a co-precipitation experiment and observation of cellular co-localization. In addition, we have obtained another TFL1 interacting protein that is isolated and kindly provided by Professor Natasha Raikhel at the University of California Riverside. We have subcloned the full-length cDNA and created the constructs for over-expression and RNAi experiments in plants. Currently, T1 generation transgenic plants are being screened. To further understand the function of FT and TFL1, we are examining global gene expression patterns in their mutants and over-expressing plants using the latest sequencing technique at the Keck Center on campus. Our bioinformatics analysis suggests that highly similar gene sets are controlled by FT and TFL1, but in an antagonistic way. Our ultimate goal is to transfer the knowledge obtained in a model species to economically important crop plants. To achieve this goal, we comparatively work on soybean, the second most produced crop in the U.S. We have collected shoot materials from soybean plants that were treated with different day-length conditions and carried out an expression analysis of key flowering genes using quantitative RT-PCR and transcriptome sequencing techniques, and are currently analyzing the data. The preliminary results suggest: 1) Soybean FT and TFL1 homologs show an oscillated mRNA expression pattern in a day under control of circadian clocks, 2) The expression of soybean CO, FT, and TFL1 homologs are controlled by day-lengths, and 3) The known flowering loci in soybean E1, E2, E3, and E5 show differential expression patterns of the FT homologs. A large set of new RNA samples have been submitted to the Keck Center for sequencing to complete this project. PARTICIPANTS: Postroctoral training opportunity was provided to Yu Mei and Valpuri Sovero. Graduate student research opportunities were provided to Waseem Haider, William Brian Price, and Eric Sedivy. Undergraduate student research opportunity was provided to Gabriela Seufferheld. Statistical research training opportunity was provided to Hongqiang Sun. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Our results and achievements obtained in 2011 became a basis of three public poster presentations in scientific conferences and two invited seminars domestic and abroad. These presentations significantly enhanced the PI's scientific interaction with the community and helped to enhance the recognition of the scholarly activities of the PI and the University in public. These activities provided the PI opportunities to develop new collaborative interactions with researchers in other institutions. In addition, we have submitted two manuscripts during 2011 and also are currently preparing 2 research papers, aiming for submission in early 2012. The scientific resources obtained through our activities using the support of the Hatch funds will further bring novel results, public presentations, and publications in 2012 and the following years. Furthermore, the material and results obtained in 2011 using the Hatch funds provided us with a foundation to develop a new program. In addition to a USDA-AFRI grant ($500K) that started in April 2011, we have applied two large federal grants (DOE: $1M, NSF: $3M) in collaboration with other faculty members on campus and three small industry based funds, although unsuccessful. We will carry on the efforts and are hopeful that our efforts will eventually succeed to obtain additional funding.
Publications
- Hanzawa, Y. 2011. Evolutionary and ecological roles of FRIGIDA in the Arabidopsis flowering gene networks. (Pending).
- Sovero, V. and Hanzawa, Y. 2011. Flowering control in short-day flowering plants. (Pending).
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: Our postdoctoral researcher Yu Mei tested our hypothesis where CONSTANS, a key gene in the photoperiodic pathways in Arabidopsis thaliana, regulates the expression of a meristem identity gene TFL1. We have been developing a collaboration with Professor Francisco Madueno in Spain and exchanged this data with his group. Currently we are working on a manuscript to publish the result, aiming for the submission in spring 2011. To identify novel function of Arabidopsis flowering gene TFL1 in plant development, we tested the effects of the loss-of-function mutant of TFL1 and over-expression of TFL1 in root gravitropism. Similarly, the root gravitropism effects of a potential TFL1 interactor TIL3 and its homologs, TIL3L1, TIL3L2, and TIL3L3, were tested. Flowering time phenotypes of the mutants and the TFL1 over-expressing plants were also measured in comparison with wild-type plants. Aiming to further characterize the protein-protein interaction of TFL1 and the TIL3 family, we have sub-cloned the full-length cDNAs of TFL1 and the TIL3 family, as well as FT, a TFL1 homolog, and created the constructs to express these proteins with a fluorescent tag in both the N-ter and the C-ter fusion forms. These constructs will be used to transform plants to test protein localization and co-localization by observing the fluorescent signals under a microscope. In addition, Yu and an undergraduate lab assistant have created double mutants of these mutations in various combinations to examine whether the double mutants show additive or non-additive phenotypic effects. To develop this project further, we have developed a collaborative interaction with Professor Natasha Raikhel at the University of California Riverside and Professor Glenda Gillaspy at Virginia Tech on this project. The phenotypic data and the constructs obtained in this project have been shared with Professors Raikhel and Gillaspy. Our ultimate goal is to transfer the knowledge obtained in a model species to economically important crop plants. To achieve this goal, we are carrying out the profiling of gene regulation patterns in soybean, the second most produced crop in the U.S., in response to a day-length change that influences flowering transition. A graduate student, Brian Price, collected shoot materials from soybean plants that were treated with a day-length shift in summer/fall 2010. To pursue this project, we applied for external funding and successfully obtained a USDA AFRI grant of $500K. We are also developing a close collaboration with Professor Randall Nelson at the USDA and Professor Matthew Hudson in the department on this project. PARTICIPANTS: This project provided a training opportunity to three lab personnel including Yu Mei, a postdoctoral researcher, and William Brian Price, a MS graduate student, and Gabriela Seufferheld, an undergraduate student, to develop ability to design, carry out laboratory experiment, and analyze biological data sets. Potential collaborators: Randall Nelson, USDA and Department of Crop Sciences, University of Illinois. Matthew Hudson, Department of Crop Sciences, University of Illinois. Michael D. Purugganan, Department of Biology, New York University. Natasha Raikhel, Center for Plant Cell Biology, University of California, Riverside. Glenda Gillaspy, Biochemistry, Virginia Tech. Francisco Madueno, University of Valencia, Spain. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Our identification of the regulatory action of CONSTANS on the expression of TFL1 brought an attractive and unique evolutionary hypothesis that a regulatory gene can maintain two opposing downstream effects. Furthermore, we obtained a piece of evidence that these opposing effects are mediated by a set of homologous genes. These results have attracted other researchers that led us to the international collaborative publication. This work addressed a part of our objective 1. In addition, we achieved a better understanding of the TFL1 pathway and the interaction with the TIL3 family in plant development that are the major questions of objective 1. The genetic resources obtained through our activities using the support of the hatch funds will further bring novel results and publications in 2011 and the following years. Furthermore, the material and results obtained in 2010 provided us a foundation to develop a new program. For the soybean project, it offered preliminary results for a successfull proposal and helped to develop internal collaborations for expansion of the project. For the Arabidopsis project, we have collected sufficient data to prepare for a publication, as well as new external collaborations. We are also planning to apply for funding in collaboration with Professor Gillaspy in 2011 based on the preliminary data that we have collected using the hatch funds in part.
Publications
- No publications reported this period
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