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ARS Home » Pacific West Area » Albany, California » Plant Gene Expression Center » Research » Research Project #425045

Research Project: Characterizing Circadian Regulatory Networks in Grain Crops to Establish their Role in Development and Abiotic Responses

Location: Plant Gene Expression Center

2017 Annual Report

The circadian clock in crop plants controls important performance traits including growth, timing of flowering and stress and pathogen responses by coordinating daily and seasonal changes in physiology. The long term goal of this project is to define at the genetic and molecular levels the circadian system in grain crops, including corn, and its impact on agronomic traits. This project will use genomic, genetic, and molecular methods to identify and characterize the circadian system in corn, utilizing resources and tools in corn and other model plant systems as appropriate. The circadian system genes identified will provide gene targets for enhancing crop performance and adaptation to global climate change. The objectives of the project are: Objective 1: Identify and characterize genes required for circadian rhythms in grain crops. Objective 2: Identify and characterize genes required for circadian clock-regulated developmental processes in grain crops. Objective 3: Analyze the contribution of the circadian system to drought responses in grain crops.

The genes required for circadian rhythms in maize remain uncharacterized. The goal of Objective 1 is to identify and/or construct mutants in candidate genes to define the genes that participate in the core circadian oscillator. Subsequent analysis of mutants will establish the function of their gene products to understand the molecular nature of the maize circadian oscillator. The hypotheses to be tested are: Mutations in circadian clock genes will alter circadian clock-driven transcription; Additional mutant alleles in clock genes can be identified and constructed using publicly available germplasm collections; and, Regional mutagenesis with the Ds transposon will create additional gi2 knockout alleles. Work in model plants demonstrates that the circadian system is deeply imbedded in regulatory networks that control growth and developmental processes. Whether such a regulatory system exists in maize remains an open question. The goal of Objective 2 is to investigate whether circadian regulation is an important contributor to maize growth and development by studying circadian clock mutants. The hypotheses to be tested are: Maize circadian clock genes are involved in regulation of maize flowering time; gi functions within the genetic networks known to control maize flowering time; The gi and tocl1 genes underlie known flowering time QTL; gi activity contributes to the timing of the juvenile to adult transition; and, Maize clock genes participate in regulation of growth. Specific core circadian oscillator genes play important roles in the responses of model plants to drought stress, in part through regulation of phytohormone signaling. The goal of Objective 3 is test whether drought stress and phytohormone responses in maize depend on the activity of circadian clock genes. The hypotheses to be tested are: The tocl1 gene is involved in maize drought responses; and, The tocl1 gene contributes to ABA responses in maize.

Progress Report
For all Objectives, the goal is to understand gene function as a means to discover how grain crops resist environmental stresses and/or control developmental transitions. This year saw progress in establishment and analysis of mutants in maize and the model plant Arabidopsis thaliana. Preliminary work was carried out to expand the scope of the study to evaluating gene function in sorghum, an important forage, grain, and biofuel crop plant closely related to maize. Regarding Objectives 1 and 2, analysis of mature plant growth for maize without the activity of the related gigantea (gi) genes, gi1 and gi2, showed that these plants grow taller than normal plants, but not any more so than mutant plants lacking a working copy of gi1 or gi2. This observation indicates that the gi1 and gi2 genes act together, instead of separately, to ensure maize plants reach an appropriate height. Genetic crossing proceeded to combine the same gi mutants together in several other diverse maize genetic backgrounds. The purpose of this effort is to learn whether the activity of these gi genes is influenced by the extensive genetic diversity present in maize, which is important both to discover new genes and to enable future efforts to use these findings for development of new maize lines. The impact of these efforts is that engineering and breeding efforts to modify maize growth could focus on differentially active versions of the gi1 and gi2 genes. For the same reasons, this information is useful for improvement of related grass species like sorghum, sugarcane, and millet. Recently ARS scientists in Lubbock, Texas publically released a large collection of mutant lines for sorghum, a close relative of maize. This collection provides the opportunity to expand the scope of Objectives 1-3 to include studies of circadian clock-associated gene activity in sorghum. A collaboration was established with the ARS scientists in Lubbock, Texas with the goal of addressing whether sorghum circadian clock-associated genes, including gi, are important for developmental processes and/or resistance to environmental stresses, including heat, cold and drought. This collaboration will also enable comparative studies of regulatory processes between maize and sorghum, which is relevant to Objectives 1 and 2. Work in the model plant Arabidopsis identified a mutant that causes substantial alterations of circadian rhythms. Additional study of this mutant revealed problems with regulatory networks the plants employ to effectively respond to stressful environments, including cold and elevated salt. These findings are relevant to the broad goal of Objective 3 to understand the role of circadian clock-associated genes in plant stress responses. The impact of work on Objectives 1-3 is a deeper understanding of how specific genes in crop plants provide the capacity for environmental adaptation through control of developmental processes and stress responses.

1. Discovery of a gene used by plants to resist the adverse consequences of low temperature. To learn how plants know growth conditions change and what actions they take to achieve their highest growth potential in these situations, ARS scientists in Albany, California studied the behavior of a well characterized plant, Arabidopsis thaliana, which is a stand-in for crop plants. Their work demonstrates that a newly discovered gene is part of the system plants use to detect and respond when their environment gets colder. Because similar genes are found in all major crop species, the knowledge gained from this study can be translated to grain and specialty crop plants. It is anticipated that this knowledge will help breeders and scientists develop crop lines that can be planted earlier in the year or to expand production to regions where low temperatures currently impede crop growth.

2. Development of an efficient, simple, and accurate method to characterize genetically transformed plants. A goal of biotechnology is to develop crop plants with superior performance, such as improved drought tolerance, increased grain production, or enhanced levels of beneficial nutrients. An important tool for biotechnology is genetic modification of plants by addition or modification of DNA with transformation. When a new type of plant is made by genetic transformation, the properties of the genetic changes caused by transformation must be known. ARS scientists in Albany, California participated in a study that produced an improved method to assess genetically transformed plants. The method is useful for several types of important crop plants, including rice, citrus, potato, maize, tomato and wheat. This new method is an advance for the entire plant research community, especially for researchers employing biotechnology to improve crop plants.

Review Publications
Ko, D., Song, Q., Rohozinski, D., Taylor, S., Juenger, T.E., Harmon, F.G., Chen, Z. 2016. Temporal shift of circadian-mediated gene expression and carbon fixation contributes to biomass heterosis in maize hybrids. PLoS Genetics. 12(7):e1006197.
Gomes, J.M., Henning, L.M., Fuganti-Pagliarini, R., Nepomuceno, A.L., Nakayama, T.J., Molinari, H.B., Basso, M.F., Harmon, F.G. 2017. Functional characterization of a putative glycine max ELF4 transgenic aradopsis and its role during flowering control. Frontiers in Plant Science. 8:618. doi:10.3389/fpls.201.00618.
Marshall, C., Tartaglio, V., Duarte, M., Harmon, F.G. 2016. The Arabidopsis sickle mutant exhibits altered circadian clock responses to cool tempatures and tempature-dependent alternative splicing. The Plant Cell. 28(10):2560-2575. doi:10.1105/tpc.16.00223.
Collier, R.A., Dasgupta, K., Xing, Y., Hernandez, B., Shao, M., Rohozinski, D., Kovak, E., Lin, J.W., De Oliveira, M., Stover, E.W., Mc Cue, K.F., Harmon, F.G., Blechl, A.E., Thomson, J.G., Thilmony, R.L. 2017. Accurate measurement of transgene copy number in crop plants using droplet digital PCR. Plant Journal. 90:1014-1025.