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. In these plants, up to thousands of individual florets are tightly packed onto a capitulum disk. While early floret development occurs continuously across capitula to generate iconic spiral phyllotaxy, during anthesis floret development occurs in discrete ring-like pseudowhorls with up to hundreds of florets undergoing simultaneous maturation. We demonstrate circadian regulation of floral organ growth and show that the effects of light on this process are time-of-day dependent. Delays in the phase of floral anthesis delay morning visits by pollinators, while disruption of circadian rhythms in floral organ development causes loss of pseudowhorl formation and large reductions in pollinator visits. We therefore show that the sunflower circadian clock acts in concert with environmental response pathways to tightly synchronize the anthesis of hundreds of florets each day, generating spatial patterns on the developing capitulum disk. This coordinated mass release of floral rewards at predictable times of day likely promotes pollinator visits and plant reproductive success.

Most organisms, from plants to insects and humans, anticipate the rise and set of the sun through an internal biological timekeeper, called the circadian clock. Plants like the common sunflower use this clock to open their flowers at dawn in time for the arrival of pollinating insects. Sunflowers are composed of many individual flowers or florets, which are arranged in spirals around a centre following an age gradient: the oldest flowers are on the outside and youngest flowers on the inside. Each day, a ring of florets of different developmental ages coordinates their opening in a specific pattern over the day. For example, petals open at dawn, pollen is presented in the morning, and stigmas, the female organs that receive pollen, unfold in the afternoon. This pattern of flowering, or floret maturation, is repeated every day for five to ten days, creating daily rhythms of flowering across the sunflower head. Previously, it was unclear how florets within each flowering ring synchronize their flowering patterns to precise times during the day. To find out more, Marshall et al. analysed time-lapse videos of sunflowers that were exposed to different day length and temperature conditions. Sunflowers opened a new floret ring every 24 hours, regardless of the length of the day. In all three day-length scenarios (short, middle, long), the development of the florets remained highly coordinated. Even flowers kept in the dark for up to four days were able to maintain the same daily growth rhythms. This persistence of daily rhythms in the absence of environmental cues suggests that the circadian clock regulates the genetic pathways that cause sunflowers to flower. However, when sunflowers whose circadian rhythms were delayed relative to the sun were placed out in a field, the sunflowers flowered later and thus attracted less pollinators. Marshall et al. show that the circadian clock is important for regulating flowering patterns in sunflowers to ensure their successful pollination. A better understanding of the interplay between pollinators, flowering plants and their environment will provide more insight into how climate change may affect pollination efficiency. By identifying the genes and pathways underlying flowering patterns, it may be possible to develop breeds that flower at the optimal times of day to promote pollination. This could help mitigate the effects of climate change and declining populations of pollinators.

An External Coincidence Model for the Lunar Cycle Reveals Circadian Phase-Dependent Moonlight Effects on Coral Spawning.

Many marine organisms synchronously spawn at specific times to ensure the success of external fertilization in the ocean. Corals are famous examples of synchronized spawning at specific lunar phases, and two distinct spawning patterns have been observed in two dominant taxa: merulinid corals spawn at regular lunar phases, several days after the full moon, whereas Acropora corals spawn at more irregular lunar phases around the full moon. Although it has been suggested that the two coral taxa have different responses to moonlight and seawater temperature, their spawning times have never been analyzed by integrating the two environmental factors, resulting in an incomplete understanding of the regulatory mechanisms of spawning. In this study, we developed a new predictive model of coral spawning days by integrating moonlight and temperature effects based on the external coincidence model for the lunar cycle. We performed model fitting using a 10-year monitoring record of coral spawning time in Taiwan. Our model successfully demonstrated the synergistic effects of moonlight and temperature on coral spawning time (days) and provided two testable hypotheses to explain the different spawning patterns regarding the preparation (maturation) process for spawning and the sensitivity to moonlight at different circadian phases: (1) Acropora corals may have an earlier onset and longer period of preparation for spawning than merulinid corals; and (2) merulinid corals may use moonlight signals near sunset, while Acropora corals may have a similar onset at approximately midnight. This is the first study to indicate the difference in circadian phase-dependent moonlight sensitivities between coral taxa, providing a basis for underlying coral spawning mechanisms for rhythmic studies.

Hello Darkness, My Old Friend: A Tutorial of Nanda-Hamner Protocols.

Plants and animals use circadian and photoperiodic timekeeping mechanisms to respond to daily and seasonal changes in light:dark and appropriately coordinate their development. Although the mechanisms that may connect the circadian and photoperiodic clock are still unclear in many species, researchers have been using Nanda-Hamner protocols for decades to elucidate how seasonal time is measured and determine whether seasonal responses have a circadian basis in a given species. In this brief tutorial we describe how to design and interpret the results of Nanda-Hamner experiments, and provide suggestions on how to use both Nanda-Hamner protocols and modern molecular experiments to better understand the mechanisms of seasonal timekeeping.

Distinct Physiological Mechanisms Induce Latitudinal and Sexual Differences in the Photoperiodic Induction of Diapause in a Fly.

Many temperate insects enter diapause (dormancy) for overwintering in response to short days (long nights). A latitudinal cline in the critical day lengths for the photoperiodic induction of diapause has been reported in various insect species. However, the physiological mechanisms underlying this cline have remained elusive. We approached this issue in the flesh fly Sarcophaga similis, in which the photoperiodic time measurement system meets the "external coincidence model." In this model, measuring day lengths depends on whether the photoinducible phase (φi), determined by a circadian clock, is exposed to light or not. First, we detected a clear latitudinal cline in the critical day lengths of flies collected from 4 localities at different latitudes. The phase positions of the φi, which can be verified by night interruption photoperiods, also showed a clear latitudinal cline. This result supports the hypothesis that the latitudinal cline in the critical day length is produced by the difference in the phase positions of the φi among different strains. A sexual difference in the critical day length for photoperiodic induction has also been detected in various species. In this study, a sexual difference in the critical day length was observed in the southern strains but there was no sexual difference in the phase positions of the φi. This result indicates that both sexes measure photoperiods in the same manner. Males are less sensitive than females to the light pulse given at the φi, suggesting a quantitative difference in the photoperiodic time measurement and counter systems. This study clearly reveals that distinct mechanisms induce latitudinal and sexual differences in the critical day length for the photoperiodic induction of diapause in a fly.

Distribution of PERIOD-immunoreactive neurons and temporal change of the immunoreactivity under long-day and short-day conditions in the larval brain of the flesh fly Sarcophaga similis.

The flesh fly Sarcophaga similis show a clear photoperiodic response; they develop into adults under long days, whereas they arrest their development at the pupal stage under short days. Although the involvement of a circadian clock in photoperiodic time measurement is suggested in this species, the anatomical location of the clock neurons responsible for the time measurement has been unknown. We detected two PERIOD-immunoreactive cell clusters in the larval brain; one cluster was located at the dorsoanterior region and the other at the medial region. We further investigated their temporal changes in PERIOD-immunoreactivity and compared their patterns under different photoperiods.

Photoperiodic flowering: time measurement mechanisms in leaves.

Many plants use information about changing day length (photoperiod) to align their flowering time with seasonal changes to increase reproductive success. A mechanism for photoperiodic time measurement is present in leaves, and the day-length-specific induction of the FLOWERING LOCUS T (FT) gene, which encodes florigen, is a major final output of the pathway. Here, we summarize the current understanding of the molecular mechanisms by which photoperiodic information is perceived in order to trigger FT expression in Arabidopsis as well as in the primary cereals wheat, barley, and rice. In these plants, the differences in photoperiod are measured by interactions between circadian-clock-regulated components, such as CONSTANS (CO), and light signaling. The interactions happen under certain day-length conditions, as previously predicted by the external coincidence model. In these plants, the coincidence mechanisms are governed by multilayered regulation with numerous conserved as well as unique regulatory components, highlighting the breadth of photoperiodic regulation across plant species.

Verification at the protein level of the PIF4-mediated external coincidence model for the temperature-adaptive photoperiodic control of plant growth in Arabidopsis thaliana.

Plant circadian clock controls a wide variety of physiological and developmental events, which include the short-days (SDs)-specific promotion of the elongation of hypocotyls during de-etiolation and also the elongation of petioles during vegetative growth. In A. thaliana, the PIF4 gene encoding a phytochrome-interacting basic helix-loop-helix (bHLH) transcription factor plays crucial roles in this photoperiodic control of plant growth. According to the proposed external coincidence model, the PIF4 gene is transcribed precociously at the end of night specifically in SDs, under which conditions the protein product is stably accumulated, while PIF4 is expressed exclusively during the daytime in long days (LDs), under which conditions the protein product is degraded by the light-activated phyB and also the residual proteins are inactivated by the DELLA family of proteins. A number of previous reports provided solid evidence to support this coincidence model mainly at the transcriptional level of the PIF 4 and PIF4-traget genes. Nevertheless, the diurnal oscillation profiles of PIF4 proteins, which were postulated to be dependent on photoperiod and ambient temperature, have not yet been demonstrated. Here we present such crucial evidence on PIF4 protein level to further support the external coincidence model underlying the temperature-adaptive photoperiodic control of plant growth in A. thaliana.