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While much is known about the genes and proteins that make up the circadian clocks in vertebrates and several arthropod species, much less is known about the clock genes in many other invertebrates, including nudibranchs. The goal of this project was to identify the RNA and protein products of putative clock genes in the central nervous system of three nudibranchs, Hermissenda crassicornis, Melibe leonina, and Tritonia diomedea. Using previously published transcriptomes (Hermissenda and Tritonia) and a new transcriptome (Melibe), we identified nudibranch orthologs for the products of five canonical clock genes: brain and muscle aryl hydrocarbon receptor nuclear translocator like protein 1, circadian locomotor output cycles kaput, non-photoreceptive cryptochrome, period, and timeless. Additionally, orthologous sequences for the products of five related genes-aryl hydrocarbon receptor nuclear translocator like, photoreceptive cryptochrome, cryptochrome DASH, 6-4 photolyase, and timeout-were determined. Phylogenetic analyses confirmed that the nudibranch proteins were most closely related to known orthologs in related invertebrates, such as oysters and annelids. In general, the nudibranch clock proteins shared greater sequence similarity with Mus musculus orthologs than Drosophila melanogaster orthologs, which is consistent with the closer phylogenetic relationships recovered between lophotrochozoan and vertebrate orthologs. The suite of clock-related genes in nudibranchs includes both photoreceptive and non-photoreceptive cryptochromes, as well as timeout and possibly timeless. Therefore, the nudibranch clock may resemble the one exhibited in mammals, or possibly even in non-drosopholid insects and oysters. The latter would be evidence supporting this as the ancestral clock for bilaterians.

Sequences of Circadian Clock Proteins in the Nudibranch Molluscs Hermissenda crassicornis, Melibe leonina, and Tritonia diomedea