Transcriptomics and metatranscriptomics in zooplankton: wave of the future?

Abstract Molecular tools have changed the understanding of zooplankton biodiversity, speciation, adaptation, population genetics and global patterns of connectivity. However, the molecular resources needed to capitalize on these advances continue to be limited in comparison with those available for other eukaryotic plankton. This deficiency could be addressed through an Ocean Zooplankton Open 'Omics Project (Ocean ZOOP) that would generate de novo assembled transcriptomes for hundreds of metazoan plankton species. A collection of comparable reference transcriptomes would generate a new framework for ecological and physiological studies. Defining species niches, identifying optimal habitats, assessing adaptive capacity and predicting changes in phenology are just a few examples of how such a resource could transform studies on zooplankton ecology

Diapause vs. reproductive programs: transcriptional phenotypes in a keystone copepod

© The Author(s), 2021. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Lenz, P. H., Roncalli, V., Cieslak, M. C., Tarrant, A. M., Castelfranco, A. M., & Hartline, D. K. Diapause vs. reproductive programs: transcriptional phenotypes in a keystone copepod. Communications Biology, 4(1), (2021): 426, https://doi.org/10.1038/s42003-021-01946-0.Many arthropods undergo a seasonal dormancy termed “diapause” to optimize timing of reproduction in highly seasonal environments. In the North Atlantic, the copepod Calanus finmarchicus completes one to three generations annually with some individuals maturing into adults, while others interrupt their development to enter diapause. It is unknown which, why and when individuals enter the diapause program. Transcriptomic data from copepods on known programs were analyzed using dimensionality reduction of gene expression and functional analyses to identify program-specific genes and biological processes. These analyses elucidated physiological differences and established protocols that distinguish between programs. Differences in gene expression were associated with maturation of individuals on the reproductive program, while those on the diapause program showed little change over time. Only two of six filters effectively separated copepods by developmental program. The first one included all genes annotated to RNA metabolism and this was confirmed using differential gene expression analysis. The second filter identified 54 differentially expressed genes that were consistently up-regulated in individuals on the diapause program in comparison with those on the reproductive program. Annotated to oogenesis, RNA metabolism and fatty acid biosynthesis, these genes are both indicators for diapause preparation and good candidates for functional studies.This work was supported by National Science Foundation Grants (NSF) OCE-1459235 and OCE-1756767 to P.H.L., D.K.H. and AE Christie and OPP-1746087 to A.M.T

Capital Breeding in a Diapausing Copepod: A Transcriptomics Analysis

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Prediction of a neuropeptidome for the eyestalk ganglia of the lobster Homarus americanus using a tissue-specific de novo assembled transcriptome

In silico transcriptome mining is a powerful tool for crustacean peptidome prediction. Using homology-based BLAST searches and a simple bioinformatics workflow, large peptidomes have recently been predicted for a variety of crustaceans, including the lobster, Homarus americanus. Interestingly, no in silico studies have been conducted on the eyestalk ganglia (lamina ganglionaris, medulla externa, medulla interna and medulla terminalis) of the lobster, although the eyestalk is the location of a major neuroendocrine complex, i.e., the X-organ-sinus gland system. Here, an H. americanus eyestalk ganglia-specific transcriptome was produced using the de novo assembler Trinity. This transcriptome was generated from 130,973,220 Illumina reads and consists of 147,542 unique contigs. Eighty-nine neuropeptide-encoding transcripts were identified from this dataset, allowing for the deduction of 62 distinct pre/preprohormones. Two hundred sixty-two neuropeptides were predicted from this set of precursors; the peptides include members of the adipokinetic hormone-corazonin-like peptide, allatostatin A, allatostatin B, allatostatin C, bursicon α, CCHamide, corazonin, crustacean cardioactive peptide, crustacean hyperglycemic hormone (CHH), CHH precursor-related peptide, diuretic hormone 31, diuretic hormone 44, eclosion hormone, elevenin, FMRFamide-like peptide, glycoprotein hormone α2, glycoprotein hormone β5, GSEFLamide, intocin, leucokinin, molt-inhibiting hormone, myosuppressin, neuroparsin, neuropeptide F, orcokinin, orcomyotropin, pigment dispersing hormone, proctolin, pyrokinin, red pigment concentrating hormone, RYamide, short neuropeptide F, SIFamide, sulfakinin, tachykinin-related peptide and trissin families. The predicted peptides expand the H. americanus eyestalk ganglia neuropeptidome approximately 7-fold, and include 78 peptides new to the lobster. The transcriptome and predicted neuropeptidome described here provide new resources for investigating peptidergic signaling within/from the lobster eyestalk ganglia

Diatom bloom-derived biotoxins cause aberrant development and gene expression in the appendicularian chordate Oikopleura dioica.

Investigating environmental hazards than could affect appendicularians is of prime ecological interest because they are among the most abundant components of the mesozooplankton. This work shows that embryo development of the appendicularian Oikopleura dioica is compromised by diatom bloom-derived biotoxins, even at concentrations in the same range as those measured after blooms. Developmental gene expression analysis of biotoxin-treated embryos uncovers an aberrant golf ball-like phenotype affecting morphogenesis, midline convergence, and tail elongation. Biotoxins induce a rapid upregulation of defensome genes, and considerable delay and silencing of zygotic transcription of developmental genes. Upon a possible future intensification of blooms associated with ocean warming and acidification, our work puts an alert on the potential impact that an increase of biotoxins may have on marine food webs, and points to defensome genes as molecular biosensors that marine ecologists could use to monitor the genetic stress of natural populations exposed to microalgal blooms

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

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

The Effect of the Toxic Dinoflagellate Alexandrium fundyense on the Calanoid Copepod Calanus finmarchicus

Ph.D. University of Hawaii at Manoa 2015.Includes bibliographical references.Harmful algal blooms have represented a serious problem for local economies and public health worldwide. The propagation of toxins through the marine web chain, and the negative effect on higher consumers, has raised questions on how primary consumers, such as herbivorous zooplankton respond to toxic algae. Inshore and offshore waters of the Gulf of Maine (USA) are seasonally dominated by the toxic dinoflagellate Alexandrium fundyense, which is responsible for paralytic shellfish poisoning (PSP) in humans. The calanoid copepod Calanus finmarchicus co-occurs with A. fundyense during spring and summer blooms. High survival has been reported for this copepod exposed to the dinoflagellate, however, little is known about other effects. The goal of my dissertation work was to study the effect of A. fundyense on the fitness and physiology of C. finmarchicus in controlled laboratory experiments. Fitness was measured as survival and reproductive success. Changes in global gene expression using RNA-Seq technology were used to measure the physiological response. The results suggest that blooms of A. fundyense are an environmental challenge for C. finmarchicus with a negative effect on copepod population growth. The negative effects included a decrease in reproduction, as well as metabolic adjustments, such as a decrease in lipid biosynthesis and protein synthesis/cell growth. My results demonstrate that transcriptomics is a tool that can be used to investigate the physiological ecology of non-model species such as copepods in order to identify biological processes affected by non-optimal environmental conditions

Gene losses did not stop the evolution of big brains.

Elephants and fruit bats have evolved large brains even though they have lost a gene that is fundamental to the supply of energy to the brain when glucose is not available

The Distribution of Ferritins in Marine Copepods

Iron is an essential element for the functioning of cellular processes. Ferritins, the major intracellular iron storage proteins, convert the free Fe2+ into the nontoxic Fe3+ which can be stored and transported where needed. To date, little is known about the iron metabolism in copepods; however, in these crustaceans, ferritins have been used as biomarkers of stress and diapause. A limiting factor of these studies has been the use of a single ferritin transcript as a biomarker. In this paper, we in silico mined the publicly available copepod transcriptomes to characterize the multiplicity of the ferritin transcripts in different orders and families. We also examined the expression of ferritin in three ecologically important copepods—Calanus finmarchicus, C. helgolandicus and Temora stylifera—during development and under stress conditions. A full-length transcript encoding ferritin heavy chain has been identified in all 27 mined transcriptomes, with 50% of the species possessing multiple transcripts. Ferritin expression increased in C. finmarchicus during the early–late development transition, and in T. stylifera females exposed to oxylipins at sea. Overall, our results suggest that copepod ferritins can be involved in iron storage, larval development and stress response, thus representing potential biomarker genes for ocean health status monitoring

Gene losses did not stop the evolution of big brains.

Elephants and fruit bats have evolved large brains even though they have lost a gene that is fundamental to the supply of energy to the brain when glucose is not available