Circadian signaling in Homarus americanus: Region-specific de novo assembled transcriptomes show that both the brain and eyestalk ganglia possess the molecular components of a putative clock system.

Authors: CHRISTIE, ANDREW - University Of Hawaii; YU, ANDY - University Of Hawaii; PASCUAL, MICAH - University Of Hawaii; RONCALLI, VITTORIA - University Of Hawaii; CIESLAK, MATTHEW - University Of Hawaii; Warner, Amanda; LAMEYER, TESS - Bowdoin College; STANHOPE, MEREDITH - Bowdoin College; DICKINSON, PATSY - Bowdoin College; Hull, Joe

Submitted to: Marine Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/12/2018
Publication Date: 4/12/2018
Citation: Christie, A.E., Yu, A., Pascual, M.G., Roncalli, V., Cieslak, M.C., Warner, A.N., Lameyer, T.J., Stanhope, M.E., Dickinson, P.S., Hull, J.J. 2018. Circadian signaling in Homarus americanus: Region-specific de novo assembled transcriptomes show that both the brain and eyestalk ganglia possess the molecular components of a putative clock system.. Marine Genomics. 40:25-44. https://doi.org/10.1016/j.margen.2018.03.002.
DOI: https://doi.org/10.1016/j.margen.2018.03.002

Interpretive Summary: Almost all organisms exhibit recurring patterns in physiology and/or behavior in relation to a 24-hr day. These circadian oscillations are typically regulated by a core group of conserved genes that function in keeping track of time. In crustaceans, however, little is known about the molecular mechanisms maintaining the circadian rhythmicity or within which tissue the central clock regulator is located. Although previous studies using the American lobster suggested the central regulator may reside in the eyestalk rather than the brain, questions remain. To better characterize the molecular basis of circadian control in the American lobster, high-throughput sequencing approaches were used to generate separate transcriptomes for two tissues thought to play a role in maintaining the daily circadian rhythm: lobster brain and eyestalk ganglia. Sequences with high similarity to known clock gene products were identified in both transcriptomic datasets and subsequently confirmed using PCR. Unexpectedly, tissue specific sequences were also identified, suggesting the presence of distinct timekeeping systems. The data generated in this study will provide a foundation for future investigations how specific behaviors in the lobster and other crustaceans maintain circadian rhythmicity.

Technical Abstract: Essentially all organisms exhibit recurring patterns of physiology/behavior that oscillate with a period of ~24-h and are synchronized to the solar day. Crustaceans are no exception, with robust circadian rhythms having been documented in many members of this arthropod subphylum. However, little is known about the molecular underpinnings of their circadian rhythmicity. Moreover, the location of the crustacean central clock has not been firmly established, although both the brain and eyestalk ganglia have been hypothesized as loci. The American lobster, Homarus americanus, is known to exhibit multiple circadian rhythms, and immunodetection data suggest that its central clock is located within the eyestalk ganglia rather than in the brain. Here, brain- and eyestalk ganglia-specific transcriptomes were generated and used to assess the presence/absence of transcripts encoding the commonly recognized protein components of arthropod circadian signaling systems in these two regions of the lobster central nervous system. Transcripts encoding putative homologs of the core clock proteins clock, cryptochrome 2, cycle, period and timeless were found in both the brain and eyestalk ganglia assemblies, as were transcripts encoding similar complements of putative clock-associated, clock input pathway and clock output pathway proteins. The presence and identity of transcripts encoding core clock proteins in both regions were confirmed using PCR. These findings suggest that both the brain and eyestalk ganglia possess all of the molecular components needed for the establishment of a circadian signaling system. Whether the brain and eyestalk clocks are independent of one another or represent a single timekeeping system remains to be determined. Interestingly, while most of the proteins deduced from the identified transcripts are shared by both the brain and eyestalk ganglia, assembly-specific isoforms were also identified, e.g., several period variants, suggesting the possibility of region-specific variation in clock function, especially if the brain and eyestalk clocks represent independent oscillators.