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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

2014 Annual Report

1a. Objectives (from AD-416):
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.

1b. Approach (from AD-416):
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.

3. Progress Report:
Substantial progress has been made for Objectives 1 and 2. For Objective 1, seeds for new transposon insertion mutants have been obtained via a Cooperative Research and Development Agreement and from the public Maize Genetics Cooperation. The new lines represent potential mutant alleles for the genes late elongated hypototyl1, late elongated hypototyl2, timing of cab-like1, timing of cab-like2, pseudo-response regulator73, early flowering3-like1, and early flowering3-like2. Scientists at the Plant Gene Expression confirmed the precise location of each insertion in the corresponding disrupted gene. Beginning this year, introgressions of each transposon are being made into several homogenous inbred backgrounds to create lines for testing. For Objective 2, the effort to make the double mutant between the gi1-m1 mutant allele and indeterminate1, a strong flowering time mutant allele, continued with backcrossing to the B73 inbred background. Also, a double mutant that combines the gi1-m1 and gi2-m1 alleles together will be available by the end of FY2014. Juvenile and adult vegetative maize plants carrying a gi1 mutant allele were found to be taller, with larger leaves, which 1) indicates gi1 regulates growth processes throughout development and 2) raises the possibility that gi mutant alleles could be useful tools to enhance biomass production in maize and other grasses. For Objective 3, the existing tocl2-m1 mutant allele was further backcrossed into the B73 and A632 inbred backgrounds to prepare for future drought tolerance work. A preliminary drought treatment experiment was done to establish the parameters, both experimentally and phenotypically, for testing the drought response of tocl2 and tocl1 mutant alleles.

4. Accomplishments
1. Drought modifies daily regulation of drought-responsive and circadian clock genes in soybean. The drought response pathways in soybean are incompletely understood. To identify genes involved in soybean drought responses and to test for feedback regulation between the circadian clock and drought stress pathways, ARS researchers from the Plant Gene Expression Center in Albany, California, and Brazilian researchers from the Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA) or the Brazilian Enterprise for Agricultural Research, compared the genome wide gene expression behavior of soybean plants exposed or not to water limitation. This study found regulatory crosstalk between the soybean circadian clock and drought stress-signaling pathway. It also discovered novel drought-responsive transcription factors. This study provides a more complete understanding of the signaling pathways for drought tolerance and new gene targets to improve drought tolerance in soybean and related species.

Review Publications
Gomes, J.M., Rodrigues, F.A., Fuganti-Pagliarini, R., Nakayama, T.J., Bendix, C., Celeya, R., Molinari, H.B., Nepomuceno, A.L., Harmon, F.G. 2014. Diurnal oscillations of soybean circadian clock and drought responsive genes. PLoS One. 9(1):e86402. doi:10.1371/journal.pone.0086402.

Thines, B.C., Youngwon, Y., Duarte, M.I., Harmon, F.G. 2014. The time of day effects of warm temperature on flowering time involve PIF4 and PIF5. Journal of Experimental Botany. 65(4):1141-51. DOI: 10.1093/jxb/ert487.