A Luciferase Imaging-Based Assay for Studying Temperature Compensation of the Circadian Clock.

The pace of circadian rhythms remains relatively unchanged across a physiologically relevant range of temperatures, a phenomenon known as temperature compensation. Temperature compensation is a defining characteristic of circadian rhythms, ensuring that clock-regulated processes occur at approximately the same time of day across a wide range of conditions. Despite the identification of several genes involved in the regulation of temperature compensation, the molecular mechanisms underlying this process are still not well understood.

Light quality-dependent roles of REVEILLE proteins in the circadian system.

Several closely related Myb-like activator proteins are known to have partially redundant functions within the plant circadian clock, but their specific roles are not well understood. To clarify the function of the , , and transcriptional activators, we characterized the growth and clock phenotypes of CRISPR-Cas9-generated single, double, and triple mutants. We found that these genes act synergistically to regulate flowering time, redundantly to regulate leaf growth, and antagonistically to regulate hypocotyl elongation.

Multiple light signaling pathways control solar tracking in sunflowers.

Sunflowers are famous for their ability to track the sun throughout the day and then reorient at night to face east the following morning. This occurs by differential growth patterns, with the east sides of stems growing more during the day and the west sides of stems growing more at night. This process, termed heliotropism, is generally believed to be a specialized form of phototropism; however, the underlying mechanism is unknown.

Myb-like transcription factors have epistatic effects on circadian clock function but additive effects on plant growth.

The functions of closely related Myb-like repressor and Myb-like activator proteins within the plant circadian oscillator have been well-studied as separate groups, but the genetic interactions between them are less clear. We hypothesized that these repressors and activators would interact additively to regulate both circadian and growth phenotypes. We used CRISPR-Cas9 to generate new mutant alleles and performed physiological and molecular characterization of plant mutants for five of these core Myb-like clock factors compared with a repressor mutant and an activator mutant.

XAP5 CIRCADIAN TIMEKEEPER regulates RNA splicing and the circadian clock by genetically separable pathways.

The circadian oscillator allows organisms to synchronize their cellular and physiological activities with diurnal environmental changes. In plants, the circadian clock is primarily composed of multiple transcriptional-translational feedback loops. Regulators of post-transcriptional events, such as precursor messenger RNAs (pre-mRNA) splicing factors, are also involved in controlling the pace of the clock.

The circadian clock controls temporal and spatial patterns of floral development in sunflower.

Biological rhythms are ubiquitous. They can be generated by circadian oscillators, which produce daily rhythms in physiology and behavior, as well as by developmental oscillators such as the segmentation clock, which periodically produces modular developmental units. Here, we show that the circadian clock controls the timing of late-stage floret development, or anthesis, in domesticated sunflowers.

XAP5 CIRCADIAN TIMEKEEPER Affects Both DNA Damage Responses and Immune Signaling in .

Numerous links have been reported between immune response and DNA damage repair pathways in both plants and animals but the precise nature of the relationship between these fundamental processes is not entirely clear. Here, we report that XAP5 CIRCADIAN TIMEKEEPER (XCT), a protein highly conserved across eukaryotes, acts as a negative regulator of immunity in and plays a positive role in responses to DNA damaging radiation. We find mutants have enhanced resistance to infection by a virulent bacterial pathogen, pv.

Flower orientation influences floral temperature, pollinator visits and plant fitness.

Effective insect pollination requires appropriate responses to internal and external environmental cues in both the plant and the pollinator. Helianthus annuus, a highly outcrossing species, is marked for its uniform eastward orientation of mature pseudanthia, or capitula. Here we investigate how this orientation affects floral microclimate and the consequent effects on plant and pollinator interactions and reproductive fitness.

A TILLING by sequencing approach to identify induced mutations in sunflower genes.

The Targeting Induced Local Lesions in Genomes (TILLING) technology is a reverse genetic strategy broadly applicable to every kind of genome and represents an attractive tool for functional genomic and agronomic applications. It consists of chemical random mutagenesis followed by high-throughput screening of point mutations in targeted genomic regions. Although multiple methods for mutation discovery in amplicons have been described, next-generation sequencing (NGS) is the tool of choice for mutation detection because it quickly allows for the analysis of a large number of amplicons.

Circadian Rhythms in Plants.

The circadian oscillator is a complex network of interconnected feedback loops that regulates a wide range of physiological processes. Indeed, variation in clock genes has been implicated in an array of plant environmental adaptations, including growth regulation, photoperiodic control of flowering, and responses to abiotic and biotic stress. Although the clock is buffered against the environment, maintaining roughly 24-h rhythms across a wide range of conditions, it can also be reset by environmental cues such as acute changes in light or temperature.

Arabidopsis JMJD5/JMJ30 Acts Independently of LUX ARRHYTHMO Within the Plant Circadian Clock to Enable Temperature Compensation.

The circadian system ensures that plants respond appropriately to environmental change by predicting regular transitions that occur during diel cycles. In order to be most useful, the circadian system needs to be compensated against daily and seasonal changes in temperature that would otherwise alter the pace of this biological oscillator. We demonstrate that an evening-phased protein, the putative histone demethylase JMJD5, contributes to temperature compensation.

Multiple feedback loops of the Arabidopsis circadian clock provide rhythmic robustness across environmental conditions.

Although circadian oscillators in diverse eukaryotes all depend on interlinked transcriptional feedback loops, specific components are not conserved across higher taxa. Moreover, the circadian network in the model plant is notably more complex than those found in animals and fungi. Here, we combine mathematical modeling and experimental approaches to investigate the functions of two classes of Myb-like transcription factors that antagonistically regulate common target genes.

Growth-mediated plant movements: hidden in plain sight.

While fast plant movements are spectacular but rare, almost all plants exhibit relatively slow, growth-mediated tropic movements that are key to their survival in the natural world. In this brief review, we discuss recent insights into the molecular mechanisms underlying phototropism, gravitropism, hydrotropism, and autostraightening. Careful molecular genetic and physiological studies have helped confirm the importance of lateral auxin gradients in gravitropic and phototropic responses.

Guidelines for Genome-Scale Analysis of Biological Rhythms.

Genome biology approaches have made enormous contributions to our understanding of biological rhythms, particularly in identifying outputs of the clock, including RNAs, proteins, and metabolites, whose abundance oscillates throughout the day. These methods hold significant promise for future discovery, particularly when combined with computational modeling. However, genome-scale experiments are costly and laborious, yielding "big data" that are conceptually and statistically difficult to analyze.

Circadian rhythms vary over the growing season and correlate with fitness components.

Circadian clocks have evolved independently in all three domains of life, suggesting that internal mechanisms of time-keeping are adaptive in contemporary populations. However, the performance consequences of either discrete or quantitative clock variation have rarely been tested in field settings. Clock sensitivity of diverse segregating lines to the environment remains uncharacterized as do the statistical genetic parameters that determine evolutionary potential.

The REVEILLE Clock Genes Inhibit Growth of Juvenile and Adult Plants by Control of Cell Size.

The circadian clock is a complex regulatory network that enhances plant growth and fitness in a constantly changing environment. In Arabidopsis (), the clock is composed of numerous regulatory feedback loops in which () and its homologs and act in a partially redundant manner to promote clock pace. Here, we report that the remaining members of the clade, and , play only minor roles in the regulation of clock function.

auxin biosynthetic genes are required for Arabidopsis shade avoidance.

Plants respond to neighbor shade by increasing stem and petiole elongation. Shade, sensed by phytochrome photoreceptors, causes stabilization of proteins and subsequent induction of auxin biosynthetic genes. To investigate the role of genes in phytochrome-mediated elongation, we examined auxin signaling kinetics after an end-of-day far-red (EOD-FR) light treatment, and found that an auxin responsive reporter is rapidly induced within 2 hours of far-red exposure.

Circadian regulation of sunflower heliotropism, floral orientation, and pollinator visits.

Young sunflower plants track the Sun from east to west during the day and then reorient during the night to face east in anticipation of dawn. In contrast, mature plants cease movement with their flower heads facing east. We show that circadian regulation of directional growth pathways accounts for both phenomena and leads to increased vegetative biomass and enhanced pollinator visits to flowers.

Circadian regulation of hormone signaling and plant physiology.

The survival and reproduction of plants depend on their ability to cope with a wide range of daily and seasonal environmental fluctuations during their life cycle. Phytohormones are plant growth regulators that are involved in almost every aspect of growth and development as well as plant adaptation to myriad abiotic and biotic conditions. The circadian clock, an endogenous and cell-autonomous biological timekeeper that produces rhythmic outputs with close to 24-h rhythms, provides an adaptive advantage by synchronizing plant physiological and metabolic processes to the external environment.

Reassess the t Test: Interact with All Your Data via ANOVA.

Plant biology is rapidly entering an era where we have the ability to conduct intricate studies that investigate how a plant interacts with the entirety of its environment. This requires complex, large studies to measure how plant genotypes simultaneously interact with a diverse array of environmental stimuli. Successful interpretation of the results from these studies requires us to transition away from the traditional standard of conducting an array of pairwise t tests toward more general linear modeling structures, such as those provided by the extendable ANOVA framework.

A Constitutively Active Allele of Phytochrome B Maintains Circadian Robustness in the Absence of Light.

The sensitivity of the circadian system to light allows entrainment of the clock, permitting coordination of plant metabolic function and flowering time across seasons. Light affects the circadian system via both photoreceptors, such as phytochromes and cryptochromes, and sugar production by photosynthesis. In the present study, we introduce a constitutively active version of phytochrome B-Y276H (YHB) into both wild-type and phytochrome null backgrounds of Arabidopsis (Arabidopsis thaliana) to distinguish the effects of photoreceptor signaling on clock function from those of photosynthesis.

Yeast X-chromosome-associated protein 5 (Xap5) functions with H2A.Z to suppress aberrant transcripts.

Chromatin regulatory proteins affect diverse developmental and environmental response pathways via their influence on nuclear processes such as the regulation of gene expression. Through a genome-wide genetic screen, we implicate a novel protein called X-chromosome-associated protein 5 (Xap5) in chromatin regulation. We show that Xap5 is a chromatin-associated protein acting in a similar manner as the histone variant H2A.

Turning heads: the biology of solar tracking in sunflower.

Solar tracking in the common sunflower, Helianthus annuus, is a dramatic example of a diurnal rhythm in plants. During the day, the shoot apex continuously reorients, following the sun's relative position so that the developing heads track from east to west. At night, the reverse happens, and the heads return and face east in anticipation of dawn.

Global profiling of the circadian transcriptome using microarrays.

Circadian rhythms are biological cycles with a period length of approximately 24 h that are generated by endogenous clocks. The application of microarrays for high-throughput transcriptome analysis has led to the insight that substantial portions of the transcriptomes of both humans and many model organisms are clock-regulated. In a typical circadian time course microarray experiment, samples are collected from organisms maintained in constant environmental conditions, gene expression at each time point is determined using microarrays, and finally clock-regulated transcripts are identified using statistical algorithms.