BackgroundCircadian rhythms regulate many physiological, behavioral and reproductive processes. These rhythms are often controlled by light, and daily cycles of solar illumination entrain many clock regulated processes. In scleractinian corals a number of different processes and behaviors are associated with specific periods of solar illumination or non-illumination—for example, skeletal deposition, feeding and both brooding and broadcast spawning.Methodology/Principal FindingsWe have undertaken an analysis of diurnal expression of the whole transcriptome and more focused studies on a number of candidate circadian genes in the coral Acropora millepora using deep RNA sequencing and quantitative PCR. Many examples of diurnal cycles of RNA abundance were identified, some of which are light responsive and damped quickly under constant darkness, for example, cryptochrome 1 and timeless, but others that continue to cycle in a robust manner when kept in constant darkness, for example, clock, cryptochrome 2, cycle and eyes absent, indicating that their transcription is regulated by an endogenous clock entrained to the light-dark cycle. Many other biological processes that varied between day and night were also identified by a clustering analysis of gene ontology annotations.Conclusions/SignificanceCorals exhibit diurnal patterns of gene expression that may participate in the regulation of circadian biological processes. Rhythmic cycles of gene expression occur under constant darkness in both populations of coral larvae that lack zooxanthellae and in individual adult tissue containing zooxanthellae, indicating that transcription is under the control of a biological clock. In addition to genes potentially involved in regulating circadian processes, many other pathways were found to display diel cycles of transcription.

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Many broadcast spawning corals in multiple reef regions release their gametes with incredible temporal precision just once per year, using the lunar cycle to set the night of spawning. Moonlight, rather than tides or other lunar-regulated processes, is thought to be the proximate factor responsible for linking the night of spawning to the phase of the Moon. We compared patterns of gene expression among colonies of the broadcast spawning coral Acropora millepora at different phases of the lunar cycle, and when they were maintained under one of three experimentally simulated lunar lighting treatments: i) lunar lighting conditions matching those on the reef, or lunar patterns mimicking either ii) constant full Moon conditions, or iii) constant new Moon conditions. Normal lunar illumination was found to shift both the level and timing of clock gene transcription cycles between new and full moons, with the peak hour of expression for a number of genes occurring earlier in the evening under a new Moon when compared to a full Moon. When the normal lunar cycle is replaced with nighttime patterns equivalent to either a full Moon or a new Moon every evening, the normal monthlong changes in the level of expression are destroyed for most genes. In combination, these results indicate that daily changes in moonlight that occur over the lunar cycle are essential for maintaining normal lunar periodicity of clock gene transcription, and this may play a role in regulating spawn timing. These data also show that low levels of light pollution may have an impact on coral biological clocks.

Little is known about how corals sense and respond to light. In this report the proteome of coral is explored using 2D protein electrophoresis in two species, Montastraea cavernosa and Acropora millepora. Multiple protein species have major shifts in abundance in both species when sampled in daylight compared to corals sampled late in the night. These changes were observed both in larvae lacking zooxanthellae and in adult tissue containing zooxanthellae, including both Pacific and Caribbean corals. When larvae kept in the dark were treated with either thapsigargin or ionomycin, compounds that raise the level of cytoplasmic calcium, the night pattern of proteins shifted to the day pattern. This implies that photoreceptors responding to light elevate calcium levels and that calcium acts as the second messenger relaying light responses in corals. Corals spawn at night, and spawning can be delayed by exposure to light or pushed forward by early artificial sunsets. In a series of behavioral experiments, treatment of corals with ionomycin or thapsigargin was found to delay broadcast spawning in M. franksi, demonstrating that pharmacologically altering cytoplasmic calcium levels generates the same response as light exposure. Together these results show that the photo-responsive cells of corals detect and respond to light by altering cytoplasmic calcium levels, similarly to the transduction pathways in complex invertebrate eyes. The primacy of cytoplasmic calcium levels in light responsivity has broad implications for coral reproduction, including predicting how different species spawn at different times after sunset and how reproductive isolation is achieved during coral speciation.