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From a Repressilator-Based Circadian Clock Mechanism to An Biosci. Biotechnol. Biochem., 77 (1), 10–16, 2013 Award Review From a Repressilator-Based Circadian Clock Mechanism to an External Coincidence Model Responsible for Photoperiod and Temperature Control of Plant Architecture in Arabodopsis thaliana Takafumi YAMASHINO Laboratory of Molecular and Functional Genomics, School of Agriculture, Nagoya University, Chikusa-ku, Nagoya 464-8601, Japan Online Publication, January 7, 2013 [doi:10.1271/bbb.120765] Circadian clocks enable organisms to define subjec- in many biological processes of almost all eukaryotic tive time, that is, to anticipate diurnal day and night organisms including plants.1) A clock provides plants cycles. Endogenous circadian rhythms regulate many with an adaptive advantage to anticipate and respond to aspects of an organism’s physiological and morpholog- daily changes in environmental conditions such as day ical growth and development. These daily oscillations and night, and seasonal changes in photoperiod.2,3) are synchronized to the environment by external cues Endogenous circadian rhythms were first studied by such as light and temperature, resulting in enhanced observation of daily leaf movements in plants. An fitness and growth vigor in plants. Recent findings insightful experimental approach to monitor biolumi- concerning biochemical properties of central oscillators nescence of transgenic plants harboring clock or clock- in Arabidopsis thaliana have advanced our understand- controlled gene promoter fusions to the firefly luciferase ing of circadian clock function. Central oscillators are gene (LUC), developed by Millar and Kay, made it composed of three classes of transcriptional repressors. possible to characterize biological rhythms more pre- The interactions among them include a repressilator cisely in a nondestructive manner.4) In a model higher structure. Output from the circadian clock is trans- plant A. thaliana, genetics, reverse genetics and bio- duced through regulating transcription of downstream chemical study uncovered various types of clock- genes directly by the oscillator components. The essen- associated genes in concert with progress in the tial role of the output pathway in the circadian system is monitoring technique of clock functions.5–8) Based on to make different elementary steps responsible for daily current knowledge of the molecular functions of clock- cellular processes exert maximum effects at specific associated genes,9) supported by modeling and simula- times of the day. Recently, significant progress was tion,10) central oscillator components to define the made in defining the mechanisms by which plant growth molecular mechanism of circadian clock are classified on a day-to-day basis is activated at specific times of the into three classes of transcriptional regulators: Myb- day in a manner dependent on photoperiod and temper- related proteins, CCA1 and LHY,11,12) pseudo-response ature conditions. Plant growth is controlled by the clock regulators with the CCT motif (PRR family), PRR9, through interactions with light and phytohormone PRR7, PRR5 and TOC1,13) and a recently identified signaling. This review focuses on the node that connects evening complex (EC) composed of LUX/PCL1 (GARP clock output to light and phytohormone signaling that family protein), ELF3 and ELF414) (Fig. 1). Transcripts coordinates plant growth with rhythmic changes in the of these clock genes are regulated in a circadian fasion. environment. CCA1 and LHY are expressed with sharp peaks in the morning.11) Circadian wave of sequential PRR9, PRR7 Key words: Arabidopsis thaliana; circadian clock; and PRR5 transcripts is observed during the day.13) The external coincidence; light signaling; EC genes (LUX, ELF3 and ELF4) and TOC1 are phytohormone signaling expressed from evening to midnight.15–18) Importantly, mutational lesions in the genes encoding these clock oscillators affect the robustness of circadian rhythms I. Transcriptional Regulation-Dependent severely. A cca1 lhy double mutant exhibits dampened Circadian Clock Mechanism in A. thaliana circadian rhythm with a short period.19) A prr9 prr7 prr5 triple mutant shows an arrhythmic circadian pheno- Circadian clocks are intrinsic, entrainable mecha- type.20,21) Loss of function mutants of any one of the nisms that generate biological rhythms with approx- genes encoding EC also result in arrhythmia.15–18) imately a 24-h period. Circadian rhythms are widespread These results suggest that each class of clock compo- This review was written in response to the author’s receipt of the Japan Society for Bioscience, Biotechnology, and Agrochemistry Award for the Encouragement of Young Scientists in 2009. Correspondence: Fax: +81-52-789-4091; E-mail: [email protected] Abbreviations: BR, brassinosteroid; ChIP-Seq, chromatin immunoprecipitation followed by deep sequencing; DELLAs, DELLA proteins; EC, evening complex; GA, gibberellic acid; R/FR, red/far-red Circadian Clock-Dependent Diurnal Control of Plant Architecture 11 Day and LHY indirectly through the direct inhibition of PRR9 PRRs, which are direct repressors of CCA1 and LHY. PRR7 TOC1 affects every class of oscillator components PRR5 directly as a transcriptional repressor (Fig. 1). In good agreement with this model, a toc1 mutant exhibits a short period and overexpression of TOC1 results in an TOC1 33,34) CCA1 LUX arrhythmic phenotype. The real biological clock ELF4 ELF3 LHY system of A. thaliana is more complicated than the Morning Evening to night simplified structure focused around the repressilator. In fact, it is suggested that direct and/or indirect transcrip- Fig. 1. A Model for Simplified Transcriptional Repression-Based tional activation function of the above-mentioned Interactions among Central Oscillators in A. thaliana. central oscillators (CCA1,35) LUX,15) ELF3,17) and ELF418,36)) and other oscillators (NOX,37) LWD1,38) nents plays a phase-specific essential role in oscillator and RVE839,40)) are incorporated into this clock network. functions, determining topological vulnerability of the Autoregulation of CCA1/LHY,11,12) the PRR family,29) clock network. Clarification of the biochemical activity and the EC genes30,36) is also suggested to be involved in of these clock components has advanced our under- the clock mechanism. Furthermore, the whole structure standing of clock mechanism. CCA1 and LHY directly the plant clock includes post-translational modification repress the expression of TOC1, LUX and ELF4 by of the oscillator components such as CK2 mediated binding specifically to a cis-element within their phosphorelation of CCA1 and LHY,25) phosphorelation promoters known as the evening element (EE: AAAAT- and ZTL mediated degradation of PRRs,41–43) and ATCT), a motif that is overrepresented in the promoters regulation of the EC by COP1 and GI,44) which is of clock-regulated evening expressed genes.15,22–25) It probably responsible for cellular localization, activity, was recently unraveled that PRR9, PRR7, and PRR5 act and/or stability of the proteins. Despite the complexity as transcriptional repressors in the clock network. They of the circadian clock mechanism, the direct transcrip- associate with CCA1 and LHY promoters in vivo and tional repression-based interactions among the central repress these genes from early daytime until the middle oscillators (CCA1/LHY, PRRs, and EC) to establish of the night.26) The molecular properties of TOC1 have the repressilator structure shown in Fig. 1 might be also been characterized, demonstrating it is a DNA- important for the Arabidopsis clock to generate robust binding transcriptional repressor of CCA1 and (self-sustaining) and entrainable biological rhythms in LHY.10,27,28) In addition to CCA1 and LHY, TOC1 binds response to external time cues such as light and to the promoters and inhibits the expression of almost all temperature. the above-mentioned central oscillator components, PRR9, PRR7, PRR5, LUX, and ELF4.27,28) Combinato- II. External Coincidence Mechanism to rial analysis of genome-wide expression study and Regulate Plant Architecture under Day chromatin immunoprecipitation followed by deep se- and Night Cycle Conditions in A. thaliana quencing (ChIP-Seq) lead to identification of direct target genes for PRR529) and TOC1.27,28) The DNA It is generally admitted that growth is controlled binding motifs enriched in these PRR family protein through interactions between light and phytohormone share weak sequence similarity, making it difficult to signaling.45,46) Photoreceptor mutants and mutants of identify the consensus sequence.27–29) This suggests that phytohormone biosynthesis or signaling exhibit abnor- the PRR family functions as transcriptional repressors in malities in photomorphogenesis processes during de- combination with other transcription factors to define the etiolation including elongation response of the embry- sequence specificity for promoter binding. In EC onic stem, the hypocotyl.47) Elongation of hypocotyls is proteins, LUX is responsible for the DNA binding also observed in shade avoidance responses under low activity for target genes. Protein binding microarray red/far-red (R/FR) ratios and/or low blue light inten- (PBM) experiments identified the consensus LUX bind- sity conditions.47) Involvement of phytohormones as ing site (LBS: GAT[A/T]CG).30) LBS is present at the well as photoreceptor mediated light signaling is promoters of PRR9, PRR7, TOC1, LUX, and ELF4.10,30) evident also in the control of growth under a canopy.48) Recent studies indicate that LUX functions as a tran-
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