CLOCK evolved in cnidaria to synchronize internal rhythms with diel environmental cues.

Abstract

The circadian clock enables anticipation of the day/night cycle in animals ranging from cnidarians to mammals. Circadian rhythms are generated through a transcription-translation feedback loop (TTFL or pacemaker) with CLOCK as a conserved positive factor in animals. However, CLOCK's functional evolutionary origin and mechanism of action in basal animals are unknown. In the cnidarian <i>Nematostella vectensis</i>, pacemaker gene transcript levels, including <i>NvClk</i> (the <i>Clock</i> ortholog), appear arrhythmic under constant darkness, questioning the role of NvCLK. Utilizing CRISPR/Cas9, we generated a <i>NvClk</i> allele mutant (<i>NvClk^Δ</i>), revealing circadian behavior loss under constant dark (DD) or light (LL), while maintaining a 24 hr rhythm under light-dark condition (LD). Transcriptomics analysis revealed distinct rhythmic genes in wild-type (WT) polypsunder LD compared to DD conditions. In LD, <i>NvClk^Δ/Δ</i> polyps exhibited comparable numbers of rhythmic genes, but were reduced in DD. Furthermore, under LD, the <i>NvClk^Δ/Δ</i> polyps showed alterations in temporal pacemaker gene expression, impacting their potential interactions. Additionally, differential expression of non-rhythmic genes associated with cell division and neuronal differentiation was observed. These findings revealed that a light-responsive pathway can partially compensate for circadian clock disruption, and that the <i>Clock</i> gene has evolved in cnidarians to synchronize rhythmic physiology and behavior with the diel rhythm of the earth's biosphere.