Introduction to the symposium—Keeping Time During Evolution : Conservation and Innovation of the Circadian Clock

Author Posting. © The Author(s), 2013. This is the author's version of the work. It is posted here by permission of Oxford University Press for personal use, not for redistribution. The definitive version was published in Integrative and Comparative Biology 53 (2013): 89-92, doi: 10.1093/icb/ict062.Diurnal and seasonal cues play critical and conserved roles in behavior, physiology, and reproduction in diverse animals. The circadian clock is a transcription-translation feedback loop that represents the molecular mechanism underlying many of these periodic processes, frequently through responses to light. Although much of the core regulatory machinery is deeply conserved among diverse animal lineages, there are also many examples of innovation in the way the clock either is constructed at the molecular-level or deployed in coordinating behavior and physiology. The nine papers contained within this issue address aspects of circadian signaling in diverse taxa, utilize wide-ranging approaches, and collectively provide thought-provoking discussion of future directions in circadian research.The symposium “Keeping Time During Animal Evolution: Conservation and Innovation of the Circadian Clock” was generously supported by the Society of Integrative and Comparative Biology and by Award 1239607 from the Integrative Organismal Systems Program at the National Science Foundation.2014-05-2

Vibrio elicits targeted transcriptional responses from copepod hosts

Author Posting. © The Author(s), 2016. This is the author's version of the work. It is posted here by permission of Federation of European Microbiological Societies for personal use, not for redistribution. The definitive version was published in FEMS Microbiology Ecology 92 (2016): fiw072, doi:10.1093/femsec/fiw072.Copepods are abundant crustaceans that harbor diverse bacterial communities, yet the nature of their interactions with microbiota are poorly understood . Here, we report that Vibrio elicits targeted transcriptional responses in the estuarine copepod Eurytemora affinis. We pre-treated E. affinis with an antibiotic-cocktail and exposed them to either a zooplankton specialist (Vibrio sp. F10 9ZB36) or a free-living species (V. ordalii 12B09) for 24 hours. We then identified via RNA-Seq a total of 78 genes that were differentially expressed following Vibrio exposure, including homologs of C-type lectins, chitin-binding proteins and saposins. The response differed between the two Vibrio treatments, with the greatest changes elicited upon inoculation with V. sp. F10. We suggest that these differentially regulated genes play important roles in cuticle integrity, the innate immune response, and general stress responses, and that their expression may enable E. affinis to recognize and regulate symbiotic vibrios. We further report that V. sp. F10 culturability is specifically altered upon colonization of E. affinis. These findings suggest that rather than acting as passive environmental vectors, copepods discriminately interact with vibrios, which may ultimately impact the abundance and activity of copepod-associated bacteria.2017-04-0

Heat shock protein expression during stress and diapause in the marine copepod Calanus finmarchicus

Author Posting. © The Author(s), 2011. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Journal of Insect Physiology 57 (2011): 665-675, doi:10.1016/j.jinsphys.2011.03.007.Calanoid copepods, such as Calanus finmarchicus, are a key component of marine food webs. C. finmarchicus undergoes a facultative diapause during juvenile development, which profoundly affects their seasonal distribution and availability to their predators. The current ignorance of how copepod diapause is regulated limits understanding of copepod population dynamics, distribution, and ecosystem interactions. Heat shock proteins (Hsps) are a superfamily of molecular chaperones characteristically upregulated in response to stress conditions and frequently associated with diapause in other taxa. In this study, 8 heat shock proteins were identified in C. finmarchicus C5 copepodids (Hsp21, Hsp22, p26, Hsp90, and 4 forms of Hsp70), and expression of these transcripts was characterized in response to handling stress and in association with diapause. Hsp21, Hsp22, and Hsp70A (cytosolic subfamily) were induced by handling stress. Expression of Hsp70A was also elevated in shallow active copepodids relative to deep diapausing copepodids, which may reflect induction of this gene by varied stressors in active animals. In contrast, expression of Hsp22 was elevated in deep diapausing animals; Hsp22 may play a role both in short-term stress responses and in protecting proteins from degradation during diapause. Expression of most of the Hsps examined did not vary in response to diapause, perhaps because the diapause of C. finmarchicus is not associated with the extreme environmental conditions (e.g., freezing, desiccation) experienced by many other taxa, such as overwintering insects or Artemia cysts.Funding for AMA was provided by the WHOI Summer Student Fellowship Program and an EPA STAR fellowship

Transcriptome-wide analysis of the response of the thecosome pteropod Clio pyramidata to short-term CO2 exposure

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 16 (2015): 1-9, doi:10.1016/j.cbd.2015.06.002.Thecosome pteropods, a group of calcifying holoplanktonic molluscs, have recently become a research focus due to their potential sensitivity to increased levels of anthropogenic dissolved CO2 in seawater and the accompanying ocean acidification. Some populations, however, already experience high CO2 in their natural distribution during diel vertical migrations. To achieve a better understanding of the mechanisms of pteropod calcification and physiological response to this sort of short duration CO2 exposure, we characterized the gene complement of Clio pyramidata, a cosmopolitan diel migratory thecosome, and investigated its transcriptomic response to experimentally manipulated CO2 conditions. Individuals were sampled from the Northwest Atlantic in the fall of 2011 and exposed to ambient conditions (~380 ppm) and elevated CO2 (~800 ppm, similar to levels experienced during a diel vertical migration) for ~10 hrs. Following this exposure the respiration rate of the individuals was measured. We then performed RNA-seq analysis, assembled the C. pyramidata transcriptome de novo, annotated the genes, and assessed the differential gene expression patterns in response to exposure to elevated CO2. Within the transcriptome, we identified homologs of genes with known roles in biomineralization in other molluscs, including perlucin, calmodulin, dermatopontin, calponin, and chitin synthases. Respiration rate was not affected by short-term exposure to CO2. Gene expression varied greatly among individuals, and comparison between treatments indicated that C. pyramidata down-regulated a small number of genes associated with aerobic metabolism and up-regulated genes that may be associated with biomineralization, particularly collagens and C- type lectins. These results provide initial insight into the effects of short term CO2 exposure on these important planktonic open-ocean calcifiers, pairing respiration rate and the gene expression level of response, and reveal candidate genes for future ecophysiological, biomaterial and phylogenetic studies.The Extreme Science and Engineering Discovery Environment (XSEDE), which is supported by National Science Foundation grant number OCI-1053575, provided computing resources for the differential expression analysis. This material is based upon work supported by the National Science Foundation’s Ocean Acidification Program under grant number OCE-1041068 (to Lawson, Wang, Lavery, and Wiebe), the Woods Hole Oceanographic Institution’s Access to the Sea program (to Tarrant, Maas and Lawson) and the WHOI postdoctoral scholarship program (to Maas)

Life cycle and early development of the thecosomatous pteropod Limacina retroversa in the Gulf of Maine, including the effect of elevated CO2 levels

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Springer for personal use, not for redistribution. The definitive version was published in Marine Biology 162 (2015): 2235-2249, doi:10.1007/s00227-015-2754-1.Thecosome pteropods are pelagic molluscs with aragonitic shells. They are considered to be especially vulnerable among plankton to ocean acidification (OA), but to recognize changes due to anthropogenic forcing a baseline understanding of their life history is needed. In the present study, adult Limacina retroversa were collected on five cruises from multiple sites in the Gulf of Maine (between 42° 22.1’–42° 0.0’ N and 69° 42.6’–70° 15.4’ W; water depths of ca. 45–260 m) from October 2013−November 2014. They were maintained in the laboratory under continuous light at 8° C. There was evidence of year-round reproduction and an individual life span in the laboratory of 6 months. Eggs laid in captivity were observed throughout development. Hatching occurred after 3 days, the veliger stage was reached after 6−7 days, and metamorphosis to the juvenile stage was after ~ 1 month. Reproductive individuals were first observed after 3 months. Calcein staining of embryos revealed calcium storage beginning in the late gastrula stage. Staining was observed in the shell gland, shell field, mantle, and shell margin in later stages. Exposure of two batches of larvae at the gastrula stage to elevated CO2 levels (800 and 1200 ppm) resulted in significantly increased mortality in comparison with individuals raised under ambient (~400 ppm) conditions and a developmental delay in the 1200 ppm treatment compared with the ambient and 800 ppm treatments.A. Thabet is grateful for a fellowship from the Egyptian Culture and Education Bureau and for mentoring from Drs. S.A. Saber, M.M. Sarhan and M.M. Fouda. Funding for this research was provided by a National Science Foundation grant to Lawson, Maas, and Tarrant (OCE-1316040). Additional support for field sampling was provided by the WHOI Coastal Ocean Institute and Pickman Foundation to Wang, Maas, and Lawson.2016-10-2

Conservation of DNA and ligand binding properties of retinoid X receptor from the placozoan Trichoplax adhaerens to human

Author Posting. © The Author(s), 2018. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in Journal of Steroid Biochemistry and Molecular Biology 184 (2018): 3-10, doi:10.1016/j.jsbmb.2018.02.010.Nuclear receptors are a superfamily of transcription factors restricted to animals. These transcription factors regulate a wide variety of genes with diverse roles in cellular homeostasis, development, and physiology. The origin and specificity of ligand binding within lineages of nuclear receptors (e.g., subfamilies) continues to be a focus of investigation geared toward understanding how the functions of these proteins were shaped over evolutionary history. Among early-diverging animal lineages, the retinoid X receptor (RXR) is first detected in the placozoan, Trichoplax adhaerens. To gain insight into RXR evolution, we characterized ligand- and DNA-binding activity of the RXR from T. adhaerens (TaRXR). Like bilaterian RXRs, TaRXR specifically bound 9-cis-retinoic acid, which is consistent with a recently published result and supports a conclusion that the ancestral RXR bound ligand. DNA binding site specificity of TaRXR was determined through protein binding microarrays (PBMs) and compared with human RXR. The binding sites for these two RXR proteins were broadly conserved (~85% shared high-affinity sequences within a targeted array), suggesting evolutionary constraint for the regulation of downstream genes. We searched for predicted binding motifs of the T. adhaerens genome within 1000 bases of annotated genes to identify potential regulatory targets. We identified 648 unique protein coding regions with predicted TaRXR binding sites that had diverse predicted functions, with enriched processes related to intracellular signal transduction and protein transport. Together, our data support hypotheses that the original RXR protein in animals bound a ligand with structural similarity to 9-cis-retinoic acid; the DNA motif recognized by RXR has changed little in more than 1 billion years of evolution; and the suite of processes regulated by this transcription factor diversified early in animal evolution.Support for AMT was provided by the Tropical Research Initiative and an Internal Research and Development Award from the Woods Hole Oceanographic Institution. AMR was supported by NIH award R15GM114740. JM was supported by NSF award 1536530 to AMR. DM-P, BF and FMS were supported by NIH award R01DK094707 to FMS

Daily cycle in oxygen consumption by the sea anemone Nematostella vectensis Stephenson

© The Author(s), 2016. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Biology Open (2016): 1-4, doi:10.1242/bio.013474.In bilaterian animals, the circadian clock is intimately involved in regulating energetic metabolism. Although cnidarians exhibit diel behavioral rhythms including cycles in locomotor activity, tentacle extension and spawning, daily cycles in cnidarian metabolism have not been described. To explore a possible circadian metabolic cycle, we maintained the anemone Nematostella vectensis in a 12 h light/dark cycle, a reversed light cycle, or in constant darkness. Oxygen consumption rates were measured at intervals using an optical oxygen meter. Respiration rates responded to entrainment with higher rates during light periods. During a second experiment with higher temporal resolution, respiration rates peaked late in the light period. The diel pattern could be detected after six days in constant darkness. Together, our results suggest that respiration rates in Nematostella exhibit a daily cycle that may be under circadian control and that the cycle in respiration rate is not driven by the previously described nocturnal increase in locomotor activity in this species.Funding was provided by the US–Israel Binational Science Foundation [Grant 201187]. I.T.J. was supported by the WHOI Summer Student Fellow program, which is partially funded by the National Science Foundation Research Experience for Undergraduates program. A.M.R. was supported by National Institutes of Heath [R15GM114740]

Ultraviolet radiation significantly enhances the molecular response to dispersant and sweet crude oil exposure in Nematostella vectensis

Author Posting. © The Author(s), 2018. This is the author's version of the work. It is posted here under a nonexclusive, irrevocable, paid-up, worldwide license granted to WHOI. It is made available for personal use, not for redistribution. The definitive version was published in Marine Environmental Research 134 (2018): 96-108, doi:10.1016/j.marenvres.2018.01.002.Estuarine organisms are subjected to combinations of anthropogenic and natural stressors, which together can reduce an organisms' ability to respond to either stress or can potentiate or synergize the cellular impacts for individual stressors. Nematostella vectensis (starlet sea anemone) is a useful model for investigating novel and evolutionarily conserved cellular and molecular responses to environmental stress. Using RNA-seq, we assessed global changes in gene expression in Nematostella in response to dispersant and/or sweet crude oil exposure alone or combined with ultraviolet radiation (UV). A total of 110 transcripts were differentially expressed by dispersant and/or crude oil exposure, primarily dominated by the down-regulation of 74 unique transcripts in the dispersant treatment. In contrast, UV exposure alone or combined with dispersant and/or oil resulted in the differential expression of 1133 transcripts, of which 436 were shared between all four treatment combinations. Most significant was the differential expression of 531 transcripts unique to one or more of the combined UV/chemical exposures. Main categories of genes affected by one or more of the treatments included enzymes involved in xenobiotic metabolism and transport, DNA repair enzymes, and general stress response genes conserved among vertebrates and invertebrates. However, the most interesting observation was the induction of several transcripts indicating de novo synthesis of mycosporine-like amino acids and other novel cellular antioxidants. Together, our data suggest that the toxicity of oil and/or dispersant and the complexity of the molecular response are significantly enhanced by UV exposure, which may co-occur for shallow water species like Nematostella.This material is based upon work supported by the National Science Foundation under Grant No. MCB1057152 (MJJ), MCB1057354 (AMT) and DEB1545539 (AMR)

Ecology and physiology of dormancy in a changing world: introduction to a virtual symposium

Author Posting. © University of Chicago, 2019. This article is posted here by permission of University of Chicago for personal use, not for redistribution. The definitive version was published in Biological Bulletin 237(2), (2019): 73-75, doi: 10.1086/706563.Dormancy is a widespread strategy used by diverse animal groups to persist through adverse environmental conditions, spread reproductive risk, and optimize seasonal phenology. Dormancy is an overarching term that refers to a reduction in metabolism, growth, and development; and different types of dormancy have been defined. Quiescence is directly initiated and terminated in response to environmental conditions, while diapause requires a preparatory phase that usually anticipates the onset of unfavorable conditions and also requires some minimum dormancy period (refractory phase) prior to termination. Dormancy is a fundamental feature of seasonal food web dynamics. Zooplankton populations can rapidly boom as individuals emerge from dormancy to feed on ephemeral algal blooms. Such productivity is critical to sustaining higher predators and supporting fisheries, particularly the growth of larval fish. Dormancy traits undergo selective pressure as zooplankton optimize developmental timing to maximize food availability and minimize predation pressure. As oceans warm and environments change, the relationship between dormancy cues, such as temperature and photoperiod, can shift, with as yet unknown effects on the timing of dormancy and resulting ecosystem dynamics. Future ecosystem dynamics are difficult to predict in part because we do not fully understand the cues that regulate the initiation or termination of dormancy, or how dormancy traits may change over time through acclimation and adaptation.2020-10-1

Exposure to CO2 influences metabolism, calcification and gene expression of the thecosome pteropod Limacina retroversa

Author Posting. © The Company of Biologists, 2018. This article is posted here by permission of The Company of Biologists for personal use, not for redistribution. The definitive version was published in Journal of Experimental Biology 221 (2018): jeb164400, doi:10.1242/jeb.164400.Thecosomatous pteropods, a group of aragonite shell-bearing zooplankton, are becoming an important sentinel organism for understanding the influence of ocean acidification on pelagic organisms. These animals show vulnerability to changing carbonate chemistry conditions, are geographically widespread, and are both biogeochemically and trophically important. The objective of this study was to determine how increasing duration and severity of CO2 treatment influence the physiology of the thecosome Limacina retroversa, integrating both gene expression and organism-level (respiration and calcification) metrics. We exposed pteropods to over-saturated, near-saturated or under-saturated conditions and sampled individuals at 1, 3, 7, 14 and 21 days of exposure to test for the effect of duration. We found that calcification was affected by borderline and under-saturated conditions by week two, while respiration appeared to be more strongly influenced by an interaction between severity and duration of exposure, showing complex changes by one week of exposure. The organismal metrics were corroborated by specific gene expression responses, with increased expression of biomineralization-associated genes in the medium and high treatments throughout and complex changes in metabolic genes corresponding to both captivity and CO2 treatment. Genes associated with other physiological processes such as lipid metabolism, neural function and ion pumping had complex responses, influenced by both duration and severity. Beyond these responses, our findings detail the captivity effects for these pelagic organisms, providing information to contextualize the conclusions of previous studies, and emphasizing a need for better culturing protocols.Funding for this research was provided by a National Science Foundation grant to G.L.L., A.E.M. and A.M.T. (OCE-1316040). Additional support for field sampling was provided by theWoods Hole Oceanographic Institution, Coastal Ocean Institute and the Pickman Foundation.2019-02-1