Phylogenetic footprint of the plant clock system in angiosperms: evolutionary processes of <Emphasis Type="Italic">Pseudo-Response Regulator</Emphasis>s

Background  

Plant circadian clocks regulate many photoperiodic and diurnal responses that are conserved among plant species. The plant circadian clock system has been uncovered in the model plant, Arabidopsis thaliana, using genetics and systems biology approaches. However, it is still not clear how the clock system had been organized in the evolutionary history of plants. We recently revealed the molecular phylogeny of LHY/CCA1 genes, one of the essential components of the clock system. The aims of this study are to reconstruct the phylogenetic relationships of angiosperm clock-associated PRR genes, the partner of the LHY/CCA1 genes, and to clarify the evolutionary history of the plant clock system in angiosperm lineages.     

Catechin promotes growth of Arabidopsis thaliana with concomitant changes in vascular system, photosynthesis and hormone content

Catechin is associated with several functions in animal and plant systems, with little information available regarding its role in plant growth. Low concentrations of catechin (50 and 100 μM) were found to enhance length of primary and lateral roots, number of lateral roots, fresh and dry masses of shoots and roots, leaf area, water potential of leaf and root tissues, the number of vascular bundles in the inflorescence, and leaf thickness in Arabidopsis thaliana ecotype Col-0. A significant increase in net photosynthetic rate, stomatal conductance and concentration of indole-3-acetic acid was also observed in catechin treated plants.     

Ectopic phytocystatin expression leads to enhanced drought stress tolerance in soybean (Glycine max) and Arabidopsis thaliana through effects on strigolactone pathways and can also result in improved seed traits

Ectopic cystatin expression has long been used in plant pest management, but the cysteine protease, targets of these inhibitors, might also have important functions in the control of plant lifespan and stress tolerance that remain poorly characterized. We therefore characterized the effects of expression of the rice cystatin, oryzacystatin‐I (OCI), on the growth, development and stress tolerance of crop (soybean) and model (Arabidopsis thaliana) plants. Ectopic OCI expression in soybean enhanced shoot branching and leaf chlorophyll accumulation at later stages of vegetative development and enhanced seed protein contents and decreased the abundance of mRNAs encoding strigolactone synthesis enzymes. The OCI‐expressing A. thaliana showed a slow‐growth phenotype, with increased leaf numbers and enhanced shoot branching at flowering. The OCI‐dependent inhibition of cysteine proteases enhanced drought tolerance in soybean and A. thaliana, photosynthetic CO₂ assimilation being much less sensitive to drought‐induced inhibition in the OCI‐expressing soybean lines. Ectopic OCI expression or treatment with the cysteine protease inhibitor E64 increased lateral root densities in A. thaliana. E64 treatment also increased lateral root densities in the max2‐1 mutants that are defective in strigolactone signalling, but not in the max3‐9 mutants that are defective in strigolactone synthesis. Taken together, these data provide evidence that OCI‐inhibited cysteine proteases participate in the control of growth and stress tolerance through effects on strigolactones. We conclude that cysteine proteases are important targets for manipulation of plant growth, development and stress tolerance, and also seed quality traits.     

What makes the Arabidopsis clock tick on time? A review on entrainment

Entrainment, the synchronization of a circadian clock with the external environment, is a crucial step in daily life. Although many signals contribute to entrainment, light and temperature are typically the strongest resetting cues. Much progress has been made concerning light resetting in the model plant Arabidopsis thaliana. Multiple photoreceptors (phytochromes, cryptochromes, LOV-domain proteins) are involved in light perception. The clock genes CCA1, LHY and TOC1 are all probable targets of light signalling, although the details of these pathways are not completely established. Temperature can entrain the clock, but little is known about the mechanism underlying this resetting; no obvious clock gene candidate for temperature resetting has been identified. Although circadian research has emphasized oscillations in free-running conditions, in the real world the circadian clock is entrained. During entrainment, short or long period mutants exhibit a 24-h period, but a mutant phenotype is often manifested as an altered phase relationship with the entraining cycle; short and long period mutants show leading and lagging phases, respectively, and this may be detrimental under some conditions. Arrhythmic CCA1-overexpressing plants display increased lethality under very short photoperiods, consistent with the circadian clock being of adaptive significance to life on a rotating world.     

Crosstalk between the Circadian Clock and Innate Immunity in Arabidopsis

The circadian clock integrates temporal information with environmental cues in regulating plant development and physiology. Recently, the circadian clock has been shown to affect plant responses to biotic cues. To further examine this role of the circadian clock, we tested disease resistance in mutants disrupted in CCA1 and LHY, which act synergistically to regulate clock activity. We found that cca1 and lhy mutants also synergistically affect basal and resistance gene-mediated defense against Pseudomonas syringae and Hyaloperonospora arabidopsidis. Disrupting the circadian clock caused by overexpression of CCA1 or LHY also resulted in severe susceptibility to P. syringae. We identified a downstream target of CCA1 and LHY, GRP7, a key constituent of a slave oscillator regulated by the circadian clock and previously shown to influence plant defense and stomatal activity. We show that the defense role of CCA1 and LHY against P. syringae is at least partially through circadian control of stomatal aperture but is independent of defense mediated by salicylic acid. Furthermore, we found defense activation by P. syringae infection and treatment with the elicitor flg22 can feedback-regulate clock activity. Together this data strongly supports a direct role of the circadian clock in defense control and reveal for the first time crosstalk between the circadian clock and plant innate immunity.     

The evening complex is central to the difference between the circadian clocks of Arabidopsis thaliana shoots and roots

The circadian clock regulates the timing of many aspects of plant physiology, and this requires entrainment of the clock to the prevailing day:night cycle. Different plant cells and tissues can oscillate with different free-running periods, so coordination of timing across the plant is crucial. Previous work showed that a major difference between the clock in mature shoots and roots involves light inputs. The objective of this work was to define, in Arabidopsis thaliana, the operation of the root clock in more detail, and in particular how it responds to light quality. Luciferase imaging was used to study the shoot and root clocks in several null mutants of clock components and in lines with aberrant expression of phytochromes. Mutations in each of the components of the evening complex (EARLY FLOWERING 3 and 4, and LUX ARRHYTHMO) were found to have specific effects on roots, by affecting either rhythmicity or period and its response to light quality. The data suggest that the evening complex is a key part of the light input mechanism that differs between shoots and roots and show that roots sense red light via phytochrome B.     

Mutations in leaf starch metabolism modulate the diurnal root growth profiles of Arabidopsis thaliana

Roots of Arabidopsis thaliana exhibit stable diurnal growth profiles that are controlled by the circadian clock. Here we describe the effects of mutations in leaf starch metabolism on the diurnal root growth characteristics of Arabidopsis thaliana. High temporal and spatial resolution video imaging was performed to quantify the growth kinetics of Arabidopsis wild-type as well as pgm, sex1, mex1, dpe1 and dpe2 starch metabolism mutants grown in three different photoperiods. As a result, root growth patterns of all genotypes displayed characteristic modifications in their diurnal kinetics that were also affected by the photoperiod. To further investigate the role of starch derived substrate deficiency on root growth, the effect of 0.05% extracellular sucrose was studied in 12 h-12 h light-dark cycles.Key words: diurnal root growth kinetics, dpe1, dpe2, mex1, pgm, sex1, starch metabolism, video imagingRoot growth of Arabidopsis thaliana is highly rhythmic with respect to the time of the day.^1–3 In general, root growth rates increase at night while most of the light period is characterized by declining elongation rates. Since a slow oscillation in root growth rate with a periodicity of approximately 24 h persists in free running conditions it was demonstrated that the circadian clock mediates these daily fluctuations.¹ Root growth at night is fueled by the degradation of starch within the leaves. Thus, a correspondence between the time taken to degrade starch reserves and the length of the night is important to optimize growth in C-limiting conditions. Gibon et al. observed a strong correlation between the rate of starch degradation and the relative growth rate when Arabidopsis Col-0 was grown in a range of different photoperiods.⁴ Therefore, to avoid periods of C starvation at the end of the night the circadian clock was postulated to function as a timer that adjusts degradation of starch to the prevailing length of the night.^1,5Root growth strongly depends on the supply of sucrose from the leaves. To investigate the effects of substrate depletion on root elongation at night, 12-day-old seedlings of Col-0, pgm and sex1 growing in a 16 h photoperiod were previously investigated by digital time resolved video imaging.¹ As a result, the diel growth response was strongly modified in pgm and sex1 as compared to the wild-type. Both mutants showed a pronounced inhibition of growth during the night and a gradual recovery of growth during the light period. To substantiate these findings, we here report on the root elongation patterns of additional mutants in starch metabolism, e.g., mex1, dpe1 and dpe2 detected at different photoperiods and elevated external sucrose supply.     

Effect of Lysophosphatidylethanolamine and Brassinosteroids on Development of <Emphasis Type="Italic">Arabidopsis</Emphasis> Roots

Exogenously applied lysophosphatidylethanolamine (LPE) increased the growth of primary roots and the formation of lateral roots in Arabidopsis thaliana. In the presence of brassinolide, lateral root formation induced by LPE was enhanced, implying that both LPE and brassinosteroids (BR) interact positively in the development of Arabidopsis roots. Co-treatment with LPE and BRs increased the bending activity in the rice lamina inclination assay compared to that when BRs were applied alone, suggesting that LPE seems to exert its activity via BRs activity. RT-PCR revealed that LPE did not alter the expressions of genes involved in the biosynthesis of BRs but did activate the expressions of BR signaling genes in A. thaliana. In a BR-insensitive mutant, bri1, enhanced gravitropic response by LPE in wild-type A. thaliana was diminished. In conclusion, LPE is a positive regulator for the growth and development of Arabidopsis roots, and this process seems to be enhanced by BR signaling rather than by increase in endogenous levels of BRs in A. thaliana.     

Circadian control of root elongation and C partitioning in Arabidopsis thaliana

Plants grow in a light/dark cycle. We have investigated how growth is buffered against the resulting changes in the carbon supply. Growth of primary roots of Arabidopsis seedlings was monitored using time‐resolved video imaging. The average daily rate of growth is increased in longer light periods or by addition of sugars. It responds slowly over days when the conditions are changed. The momentary rate of growth exhibits a robust diel oscillation with a minimum 8–9 h after dawn and a maximum towards the end of the night. Analyses with starch metabolism mutants show that starch turnover is required to maintain growth at night. A carbon shortfall leads to an inhibition of growth, which is not immediately reversed when carbon becomes available again. The diel oscillation persists in continuous light and is strongly modified in clock mutants. Central clock functions that depend on CCA1/LHY are required to set an appropriate rate of starch degradation and maintain a supply of carbon to support growth through to dawn, whereas ELF3 acts to decrease growth in the light period and promote growth in the night. Thus, while the overall growth rate depends on the carbon supply, the clock orchestrates diurnal carbon allocation and growth.     

Morphogenetic Effect of the Herbicide Cinch on Arabidopsis thaliana Root Development

Cinch is a morphogenetically active herbicide that inhibits primary root growth and induces abnormal ``nodule-like' lateral roots on Arabidopsis thaliana seedlings. Using 200 nm Cinch, the early stages of lateral root formation occurred along the apical half of the root axis; but once emerged, they were inhibited from further growth. Second-order lateral roots formed at the base of stunted first-order lateral roots after 5 days of Cinch treatment. Results from Cinch experiments suggested that pericycle cells are determined in the meristem to be potential sites of lateral root formation, and the developmental transition point between emerged lateral roots and subsequent growth is inhibited. Results using 2,4-dichlorophenoxyacetic acid and 2,3,5-triiodobenzoic acid suggest that Cinch is not a chemical analog of auxin. Received August 8, 1997; accepted February 23, 1998