Cellular remodeling and JAK inhibition promote zygotic gene expression in the Ciona germline.

Transcription control is a major determinant of cell fate decisions in somatic tissues. By contrast, early germline fate specification in numerous vertebrate and invertebrate species relies extensively on RNA-level regulation, exerted on asymmetrically inherited maternal supplies, with little-to-no zygotic transcription. However delayed, a maternal-to-zygotic transition is nevertheless poised to complete the deployment of pre-gametic programs in the germline. Here, we focus on early germline specification in the tunicate Ciona to study zygotic genome activation. We first demonstrate that a peculiar cellular remodeling event excludes localized postplasmic Pem-1 mRNA, which encodes the general inhibitor of transcription. Subsequently, zygotic transcription begins in Pem-1-negative primordial germ cells (PGCs), as revealed by histochemical detection of elongating RNA Polymerase II, and nascent Mef2 transcripts. In addition, we uncover a provisional antagonism between JAK and MEK/BMPRI/GSK3 signaling, which controls the onset of zygotic gene expression, following cellular remodeling of PGCs. We propose a 2-step model for the onset of zygotic transcription in the Ciona germline and discuss the significance of germ plasm dislocation and remodeling in the context of developmental fate specification.

Admp regulates tail bending by controlling ventral epidermal cell polarity via phosphorylated myosin localization in Ciona.

Ventral tail bending, which is transient but pronounced, is found in many chordate embryos and constitutes an interesting model of how tissue interactions control embryo shape. Here, we identify one key upstream regulator of ventral tail bending in embryos of the ascidian Ciona. We show that during the early tailbud stages, ventral epidermal cells exhibit a boat-shaped morphology (boat cell) with a narrow apical surface where phosphorylated myosin light chain (pMLC) accumulates. We further show that interfering with the function of the BMP ligand Admp led to pMLC localizing to the basal instead of the apical side of ventral epidermal cells and a reduced number of boat cells. Finally, we show that cutting ventral epidermal midline cells at their apex using an ultraviolet laser relaxed ventral tail bending. Based on these results, we propose a previously unreported function for Admp in localizing pMLC to the apical side of ventral epidermal cells, which causes the tail to bend ventrally by resisting antero-posterior notochord extension at the ventral side of the tail.

Pulsation waves along the Ciona heart tube reverse by bimodal rhythms expressed by a remote pair of pacemakers.

The heart of ascidians (marine invertebrate chordates) has a tubular structure, and heartbeats propagate from one end to the other. The direction of pulsation waves intermittently reverses in the heart of ascidians and their relatives; however, the underlying mechanisms remain unclear. We herein performed a series of experiments to characterize the pacemaker systems in isolated hearts and their fragments, and applied a mathematical model to examine the conditions leading to heart reversals. The isolated heart of Ciona robusta autonomously generated pulsation waves at ∼20 to 25 beats min-1 with reversals at ∼1 to 10 min intervals. Experimental bisections of isolated hearts revealed that independent pacemakers resided on each side and also that their beating frequencies periodically changed as they expressed bimodal rhythms, which comprised an ∼1.25 to 5.5 min acceleration/deceleration cycle of a beating rate of between 0 and 25 beats min-1. Only fragments including 5% or shorter terminal regions of the heart tube maintained autonomous pulsation rhythms, whereas other regions did not. Our mathematical model, based on FitzHugh-Nagumo equations applied to a one-dimensional alignment of cells, demonstrated that the difference between frequencies expressed by the two independent terminal pacemakers determined the direction of propagated waves. Changes in the statuses of terminal pacemakers between the excitatory and oscillatory modes as well as in their endogenous oscillation frequencies were sufficient to lead to heart reversals. These results suggest that the directions of pulsation waves in the Ciona heart reverse according to the changing rhythms independently expressed by remotely coupled terminal pacemakers.

Marine-Derived Peptides with Anti-Hypertensive Properties: Prospects for Pharmaceuticals, Supplements, and Functional Food.

Hypertension, a major health concern linked to heart disease and premature mortality, has prompted a search for alternative treatments due to side effects of existing medications. Sustainable harvesting of low-trophic marine organisms not only enhances food security but also provides a variety of bioactive molecules, including peptides. Despite comprising only a fraction of active natural compounds, peptides are ideal for drug development due to their size, stability, and resistance to degradation. Our review evaluates the anti-hypertensive properties of peptides and proteins derived from selected marine invertebrate phyla, examining the various methodologies used and their application in pharmaceuticals, supplements, and functional food. A considerable body of research exists on the anti-hypertensive effects of certain marine invertebrates, yet many species remain unexamined. The array of assessments methods, particularly for ACE inhibition, complicates the comparison of results. The dominance of in vitro and animal in vivo studies indicates a need for more clinical research in order to transition peptides into pharmaceuticals. Our findings lay the groundwork for further exploration of these promising marine invertebrates, emphasizing the need to balance scientific discovery and marine conservation for sustainable resource use.

Geography and developmental plasticity shape post-larval thermal tolerance in the golden star tunicate, Botryllus schlosseri.

Local adaptation and phenotypic plasticity play key roles in mediating organisms' ability to respond to spatiotemporal variation in temperature. These two processes often act together to generate latitudinal or elevational clines in acute temperature tolerance. Phenotypic plasticity is also subject to local adaptation, with the expectation that populations inhabiting more variable environments should exhibit greater phenotypic plasticity of thermal tolerance. Here we examine the potential for local adaptation and developmental plasticity of thermal tolerance in the widespread invasive tunicate Botryllus schlosseri. By comparing five populations across a thermal gradient spanning 4.4° of latitude in the northwest Atlantic, we demonstrate that warmer populations south of the Gulf of Maine exhibit significantly increased (∼0.2 °C) post-larval temperature tolerance relative to the colder populations within it. We also show that B. schlosseri post-larvae possess a high degree of developmental plasticity for this trait, shifting their median temperature of survival (LT50) upwards by as much as 0.18 °C per 1 °C increase in environmental temperature. Lastly, we found that populations vary in their degrees of developmental plasticity, with populations that experience more pronounced short-term temperature variability exhibiting greater developmental plasticity, suggesting the local adaptation of developmental plasticity. By comparing the thermal tolerance of populations across space and through time, we demonstrate how geography and developmental plasticity have shaped thermal tolerance in B. schlosseri. These results help inform our understanding of how species are able to adjust their thermal physiology in new environments, including those encountered during invasion and under increasingly novel climate conditions.

Distribution changes of non-self-test cells and self-tunic cells surrounding the outer body during Ciona metamorphosis.

BACKGROUND: Ascidians significantly change their body structure through metamorphosis, but the spatio-temporal cell dynamics in the early metamorphosis stage has not been clarified. A natural Ciona embryo is surrounded by maternally derived non-self-test cells before metamorphosis. However, after metamorphosis, the juvenile is surrounded by self-tunic cells derived from mesenchymal cell lineages. Both test cells and tunic cells are thought to be changed their distributions during metamorphosis, but the precise timing is unknown. RESULTS: Using a metamorphosis induction by mechanical stimulation, we investigated the dynamics of mesenchymal cells during metamorphosis in a precise time course. After the stimulation, two-round Ca2+ transients were observed. Migrating mesenchymal cells came out through the epidermis within 10 min after the second phase. We named this event "cell extravasation." The cell extravasation occurred at the same time as the backward movement of posterior trunk epidermal cells. Timelapse imaging of transgenic-line larva revealed that non-self-test cells and self-tunic cells temporarily coexist outside the body until the test cells are eliminated. At the juvenile stage, only extravasated self-tunic cells remained outside the body. CONCLUSIONS: We found that mesenchymal cells extravasated following two-round Ca2+ transients, and distributions of test cells and tunic cells changed in the outer body after tail regression.

Development and Validation of a Cyber-Physical System Leveraging EFDPN for Enhanced WSN-IoT Network Security.

In the current digital era, Wireless Sensor Networks (WSNs) and the Internet of Things (IoT) are evolving, transforming human experiences by creating an interconnected environment. However, ensuring the security of WSN-IoT networks remains a significant hurdle, as existing security models are plagued with issues like prolonged training durations and complex classification processes. In this study, a robust cyber-physical system based on the Emphatic Farmland Fertility Integrated Deep Perceptron Network (EFDPN) is proposed to enhance the security of WSN-IoT. This initiative introduces the Farmland Fertility Feature Selection (F3S) technique to alleviate the computational complexity of identifying and classifying attacks. Additionally, this research leverages the Deep Perceptron Network (DPN) classification algorithm for accurate intrusion classification, achieving impressive performance metrics. In the classification phase, the Tunicate Swarm Optimization (TSO) model is employed to improve the sigmoid transformation function, thereby enhancing prediction accuracy. This study demonstrates the development of an EFDPN-based system designed to safeguard WSN-IoT networks. It showcases how the DPN classification technique, in conjunction with the TSO model, significantly improves classification performance. In this research, we employed well-known cyber-attack datasets to validate its effectiveness, revealing its superiority over traditional intrusion detection methods, particularly in achieving higher F1-score values. The incorporation of the F3S algorithm plays a pivotal role in this framework by eliminating irrelevant features, leading to enhanced prediction accuracy for the classifier, marking a substantial stride in fortifying WSN-IoT network security. This research presents a promising approach to enhancing the security and resilience of interconnected cyber-physical systems in the evolving landscape of WSN-IoT networks.

Micropatterned Hydrogels with Highly Ordered Cellulose Nanocrystals for Visually Monitoring Cardiomyocytes.

Cardiac microphysiological systems are accurate in vitro platforms that reveal the biological mechanisms underlying cardiopathy, accelerating pharmaceutical research in this field. Current cardiac microphysiological devices and organs-on-chips consist of several layers prepared with complex, multi-step processes. Incorporating inorganic photonic crystals may cause long-term biocompatibility issues. Herein, micropatterned hydrogels with anisotropic structural colors are prepared by locking shear-oriented tunicate cellulose nanocrystals (TCNCs) in hydrogel networks through in situ polymerization, allowing the visualization and monitoring of cardiomyocytes. The anisotropic hydrogels are composed of highly ordered TCNCs with bright interference color and micro-grooved methacrylated gelatin with excellent biocompatibility. The microgroove patterns induce cardiomyocyte alignment and the autonomous beating of cardiomyocytes causes the hydrogels to deform, dynamically shifting the interference color. These micropatterned hydrogels could noninvasively monitor real-time changes of cardiomyocytes under pharmaceutical treatment and electrical stimulation through wavelength shifts in the transmittance spectra. This system provides a new way to detect the beat rate of cardiac tissue and it may contribute to high throughput develop.

The trunk-tail junctional region in Ciona larvae autonomously expresses tail-beating bursts at ∼20 second intervals.

Swimming locomotion in aquatic vertebrates, such as fish and tadpoles, is expressed through neuron networks in the spinal cord. These networks are arranged in parallel, ubiquitously distributed and mutually coupled along the spinal cord to express undulation patterns accommodated to various inputs into the networks. While these systems have been widely studied in vertebrate swimmers, their evolutionary origin along the chordate phylogeny remains unclear. Ascidians, representing a sister group of vertebrates, give rise to tadpole larvae that swim freely in seawater. In the present study, we examined the locomotor ability of the anterior and posterior body fragments of larvae of the ascidian Ciona that had been cut at an arbitrary position. Examination of more than 200 fragments revealed a necessary and sufficient body region that spanned only ∼10% of the body length and included the trunk-tail junction. 'Mid-piece' body fragments, which included the trunk-tail junctional region, but excluded most of the anterior trunk and posterior tail, autonomously expressed periodic tail-beating bursts at ∼20 s intervals. We compared the durations and intervals of tail-beating bursts expressed by mid-piece fragments, and also by whole larvae under different sensory conditions. The results suggest that body parts outside the mid-piece effect shortening of swimming intervals, particularly in the dark, and vary the burst duration. We propose that Ciona larvae express swimming behaviors by modifying autonomous and periodic locomotor drives that operate locally in the trunk-tail junctional region.

Contributions from both the brain and the vascular network guide behavior in the colonial tunicate Botryllus schlosseri.

We studied the function, development and aging of the adult nervous system in the colonial tunicate Botryllus schlosseri. Adults, termed zooids, are filter-feeding individuals. Sister zooids group together to form modules, and modules, in turn, are linked by a shared vascular network to form a well-integrated colony. Zooids undergo a weekly cycle of regression and renewal during which mature zooids are replaced by developing buds. The zooid brain matures and degenerates on this 7-day cycle. We used focal extracellular recording and video imaging to explore brain activity in the context of development and degeneration and to examine the contributions of the nervous system and vascular network to behavior. Recordings from the brain revealed complex firing patterns arising both spontaneously and in response to stimulation. Neural activity increases as the brain matures and declines thereafter. Motor behavior follows the identical time course. The behavior of each zooid is guided predominantly by its individual brain, but sister zooids can also exhibit synchronous motor behavior. The vascular network also generates action potentials that are largely independent of neural activity. In addition, the entire vascular network undergoes slow rhythmic contractions that appear to arise from processes endogenous to vascular epithelial cells. We found that neurons in the brain and cells of the vascular network both express multiple genes for voltage-gated Na+ and Ca2+ ion channels homologous (based on sequence) to mammalian ion channel genes.

(Epi)genomic adaptation driven by fine geographical scale environmental heterogeneity after recent biological invasions.

Elucidating processes and mechanisms involved in rapid local adaptation to varied environments is a poorly understood but crucial component in management of invasive species. Recent studies have proposed that genetic and epigenetic variation could both contribute to ecological adaptation, yet it remains unclear on the interplay between these two components underpinning rapid adaptation in wild animal populations. To assess their respective contributions to local adaptation, we explored epigenomic and genomic responses to environmental heterogeneity in eight recently colonized ascidian (Ciona intestinalis) populations at a relatively fine geographical scale. Based on MethylRADseq data, we detected strong patterns of local environment-driven DNA methylation divergence among populations, significant epigenetic isolation by environment (IBE), and a large number of local environment-associated epigenetic loci. Meanwhile, multiple genetic analyses based on single nucleotide polymorphisms (SNPs) showed genomic footprints of divergent selection. In addition, for five genetically similar populations, we detected significant methylation divergence and local environment-driven methylation patterns, indicating the strong effects of local environments on epigenetic variation. From a functional perspective, a majority of functional genes, Gene Ontology (GO) terms, and biological pathways were largely specific to one of these two types of variation, suggesting partial independence between epigenetic and genetic adaptation. The methylation quantitative trait loci (mQTL) analysis showed that the genetic variation explained only 18.67% of methylation variation, further confirming the autonomous relationship between these two types of variation. Altogether, we highlight the complementary interplay of genetic and epigenetic variation involved in local adaptation, which may jointly promote populations' rapid adaptive capacity and successful invasions in different environments. The findings here provide valuable insights into interactions between invaders and local environments to allow invasive species to rapidly spread, thus contributing to better prediction of invasion success and development of management strategies.

Asperopiperazines A and B: Antimicrobial and Cytotoxic Dipeptides from a Tunicate-Derived Fungus Aspergillus sp. DY001.

Investigation of the cytotoxic fractions of the ethyl acetate extract of the fermentation broth of the tunicate-derived Aspergillus sp. DY001 afforded two new dipeptides, asperopiperazines A and B (1 and 2), along with the previously reported compounds (+)-citreoisocoumarin (3) and (-)-6,8-di-O-methylcitreoisocoumarin (4). Analyses of the 1D and 2D NMR spectroscopic data of the compounds supported their structural assignments. Asperopiperazine A (1) is a cyclic dipeptide of leucine and phenylalanine moieties, which are substituted with an N-methyl and an N-acetyl group, respectively. On the other hand, asperopiperazine B (2) is a cyclic dipeptide of proline and phenylalanine moieties with a hydroxyl group at C-2 of the proline part. The absolute configuration of the amino acid moieties in 1 and 2 were determined by Marfey's analyses and DFT NMR chemical shift calculations, leading to their assignment as cyclo(l-NMe-Leu-l-NAc-Phe) and cyclo(d-6-OH-Pro-l-Phe), respectively. Asperopiperazines A and B displayed higher antimicrobial effects against Escherichia coli and Staphylococcus aureus than Candida albicans. Furthermore, compounds 1-4 displayed variable growth inhibitory effects towards HCT 116 and MDA-MB-231 cells, with asperopiperazine A as the most active one towards HCT 116.

The first record of egg masses in tunicates deposited by the snubnose sculpin, Orthonopias triacis, from the Northeastern Pacific: evidence for convergent evolution of an unusual reproductive strategy.

In this study, the authors report the first record of egg masses deposited in solitary tunicates by the snubnose sculpin, Orthonopias triacis, from the Northeastern Pacific. Four egg masses were discovered in the tunicate Ascidia ceratodes that were genetically determined to be O. triacis. Female O. triacis had long ovipositors that allow deposition of their eggs inside the atrium of the tunicates. A comparison of host-tunicate size with ovipositor length of sculpins from the Northwestern Pacific, including the genera Furcina and Pseudoblennius, revealed that O. triacis had shorter ovipositors and spawned in the atrium of smaller species of tunicates. Ancestral state reconstruction of egg deposition in solitary tunicates using 1.86Mbp RNAseq data of 20 sculpin species from Northeastern and Northwestern Pacific revealed that this unusual spawning behaviour may have evolved convergently in different species occurring in the Northeastern vs. the Northwestern Pacific.

Different strategies for tissue scaling in dwarf tailbud embryos revealed by single-cell analysis.

The tailbud stage is part of the organogenesis period-an evolutionarily conserved developmental period among chordates that is essential for determining the characteristics of the chordate body plan. When the volume of the egg is artificially decreased by cutting, ascidians produce a normal-looking but miniature (dwarf) tailbud embryo. Although cell lineages during ascidian embryogenesis are invariant, the number of cell divisions in the dwarf embryo is altered by a different mechanism in each tissue (Yamada and Nishida, 1999). Here, to elucidate the size-regulation strategies of the Ciona robusta dwarf tailbud embryo, we compared anatomical structure, developmental speed, and cell number/volume in each tissue between dwarf and wild type (WT) embryos. To do this, we constructed a 3D virtual mid-tailbud embryo (Nakamura et al., 2012). We could make a Ciona dwarf tailbud embryo from eggs with a diameter over 108 â€‹µm (correspond to â€‹> â€‹40% of the wild type egg volume). The timings of cleavage (~St. 12) and subsequent morphogenesis were nearly the same but blastomeres of animal hemisphere slightly delayed the timing of mitosis in the early cleavage period. Intriguingly, the tissue-to-tissue volume ratios of dwarf tailbud embryos were similar to those of wild type embryos suggesting that the ratio of tissue volumes is essential for maintaining the proper shape of the tailbud embryo. The number of cells in the epidermis, nervous system, and mesenchyme was significantly reduced in the dwarf embryos whereas the cell volume distribution of these tissues was similar in the dwarf and wild type. In contrast, the number of cells in the notochord, muscle, heart, and endoderm were maintained in the dwarf embryos; cell volumes were significantly reduced. Neither parameter changed in germline precursors. These results indicate that each tissue uses different scaling strategies to coordinate cell number and cell volume in accordance with the embryo size.

Two-Round Ca2+ transient in papillae by mechanical stimulation induces metamorphosis in the ascidian Ciona intestinalis type A.

Marine invertebrate larvae are known to begin metamorphosis in response to environmentally derived cues. However, little is known about the relationships between the perception of such cues and internal signalling for metamorphosis. To elucidate the mechanism underlying the initiation of metamorphosis in the ascidian, Ciona intestinalis type A (Ciona robusta), we artificially induced ascidian metamorphosis and investigated Ca2+ dynamics from pre- to post-metamorphosis. Ca2+ transients were observed and consisted of two temporally distinct phases with different durations before tail regression which is the early event of metamorphosis. In the first phase, Phase I, the Ca2+ transient in the papillae (adhesive organ of the anterior trunk) was coupled with the Ca2+ transient in dorsally localized cells and endoderm cells just after mechanical stimulation. The Ca2+ transients in Phase I were also observed when applying only short stimulation. In the second phase, Phase II, the Ca2+ transient in papillae was observed again and lasted for approximately 5-11 min just after the Ca2+ transient in Phase I continued for a few minutes. The impaired papillae by Foxg-knockdown failed to induce the second Ca2+ transient in Phase II and tail regression. In Phase II, a wave-like Ca2+ propagation was also observed across the entire epidermis. Our results indicate that the papillae sense a mechanical cue and two-round Ca2+ transients in papillae transmits the internal metamorphic signals to different tissues, which subsequently induces tail regression. Our study will help elucidate the internal mechanism of metamorphosis in marine invertebrate larvae in response to environmental cues.

Microbiome Variability across the Native and Invasive Ranges of the Ascidian Clavelina oblonga.

Ascidians are prolific colonizers of new environments and possess a range of well-studied features that contribute to their successful spread, but the role of their symbiotic microbial communities in their long-term establishment is mostly unknown. In this study, we utilized next-generation amplicon sequencing to provide a comprehensive description of the microbiome in the colonial ascidian Clavelina oblonga and examined differences in the composition, diversity, and structure of symbiont communities in the host's native and invasive ranges. To identify host haplotypes, we sequenced a fragment of the mitochondrial gene cytochrome c oxidase subunit I (COI). C. oblonga harbored a diverse microbiome spanning 42 bacterial and three archaeal phyla. Colonies in the invasive range hosted significantly less diverse symbiont communities and exhibited lower COI haplotype diversity than colonies in the native range. Differences in microbiome structure were also detected across colonies in the native and invasive range, driven largely by novel bacteria representing symbiont lineages with putative roles in nitrogen cycling. Variability in symbiont composition was also observed among sites within each range. Together, these data suggest that C. oblonga hosts a dynamic microbiome resulting from (i) reductions in symbiont diversity due to founder effects in host populations and (ii) environmental selection of symbiont taxa in response to new habitats within a range. Further investigation is required to document the mechanisms behind these changes and to determine how changes in microbiome structure relate to holobiont function and the successful establishment of C. oblonga worldwide.IMPORTANCE Nonnative species destabilize coastal ecosystems and microbial symbionts may facilitate their spread by enhancing host survival and fitness. However, we know little of the microorganisms that live inside invasive species and whether they change as the host spreads to new areas. In this study, we investigated the microbial communities of an introduced ascidian (Clavelina oblonga) and tracked symbiont changes across locations within the host's native and invasive ranges. Ascidians in the invasive range had less-diverse microbiomes, as well as lower host haplotype diversity, suggesting that specific colonies reach new locations and carry select symbionts from native populations (i.e., founder effects). Further, ascidians in the invasive range hosted a different composition of symbionts, including microbes with the potential to aid in processes related to invasion success (e.g., nutrient cycling). We conclude that the putative functionality and observed flexibility of this introduced ascidian microbiome may represent an underappreciated factor in the successful establishment of nonnative species in new environments.

Antibacterial Bicyclic Fatty Acids from a Korean Colonial Tunicate Didemnum sp.

Five new bicyclic carboxylic acids were obtained by antibacterial activity-guided isolation from a Korean colonial tunicate Didemnum sp. Their structures were elucidated by the interpretation of NMR, MS and CD spectroscopic data. They all belong to the class of aplidic acids. Three of them were amide derivatives (1-3), and the other two were dicarboxylic derivatives (4 and 5). The absolute configurations were determined by a bisignate pattern of CD spectroscopy, which revealed that the absolute configurations of amides were opposite to those of dicarboxylates at every stereogenic centers. Compound 2 exhibited the most potent antibacterial activity (MIC, 2 µg/mL).

Gap junction-dependent coordination of intercellular calcium signalling in the developing appendicularian tunicate Oikopleura dioica.

We characterized spontaneous Ca2+ signals in Oikopleura dioica embryos from pre-fertilization to gastrula stages following injection of GCaMP6 mRNA into unfertilized eggs. The unfertilized egg exhibited regular, transient elevations in intracellular Ca2+ concentration with an average duration of 4-6 s and an average frequency of about 1 every 2.5 min. Fertilization was accompanied by a longer Ca2+ transient that lasted several minutes. Thereafter, regular Ca2+ transients were reinstated that spread within seconds among blastomeres and gradually increased in duration (by about 50%) and decreased in frequency (by about 20%) by gastrulation. Peak amplitudes also exhibited a dynamic, with a transitory drop occurring at about the 4-cell stage and a subsequent rise. Each peak was preceded by about 15 s by a smaller and shorter Ca2+ increase (about 5% of the main peak amplitude, average duration 3 s), which we term the "minipeak". By gastrulation, Ca2+ transients exhibited a stereotyped initiation site on either side of the 32-64-cell embryo, likely in the nascent muscle precursor cells, and spread thereafter symmetrically in a stereotyped spatial pattern that engaged blastomeres giving rise to all the major tissue lineages. The rapid spread of the transients relative to the intertransient interval created a coordinated wave that, on a coarse time scale, could be considered an approximate synchronization. Treatment with the divalent cations Ni2+ or Cd2+ gradually diminished peak amplitudes, had only moderate effects on wave frequency, but markedly disrupted wave synchronization and normal development. The T-type Ca2+ channel blocker mibefradil similarly disrupted normal development, and eliminated the minipeaks, but did not affect wave synchronization. To assess the role of gap junctions in calcium wave spread and coordination, we first characterized the expression of two Oikopleura connexins, Od-CxA and Od-CxB, both of which are expressed during pre-gastrulation and gastrula stages, and then co-injected double-stranded inhibitory RNAs together with CGaMP6 to suppress connexin expression. Connexin mRNA knockdown led to a gradual increase in Ca2+ transient peak width, a decrease of interpeak interval and a marked disruption of wave synchronization. As seen with divalent cations and mibefradil, this desynchronization was accompanied by a disruption of normal development.

Secondary Metabolites of the Genus Didemnum: A Comprehensive Review of Chemical Diversity and Pharmacological Properties.

Tunicates (ascidians) are common marine invertebrates that are an exceptionally important source of natural products with biomedical and pharmaceutical applications, including compounds that are used clinically in cancers. Among tunicates, the genus Didemnum is important because it includes the most species, and it belongs to the most speciose family (Didemnidae). The genus Didemnum includes the species D. molle, D. chartaceum, D. albopunctatum, and D. obscurum, as well as others, which are well known for their chemically diverse secondary metabolites. To date, investigators have reported secondary metabolites, usually including bioactivity data, for at least 69 members of the genus Didemnum, leading to isolation of 212 compounds. Many of these compounds exhibit valuable biological activities in assays targeting cancers, bacteria, fungi, viruses, protozoans, and the central nervous system. This review highlights compounds isolated from genus Didemnum through December 2019. Chemical diversity, pharmacological activities, geographical locations, and applied chemical methods are described.

Diversity, Bioactivity Profiling and Untargeted Metabolomics of the Cultivable Gut Microbiota of Ciona intestinalis.

It is widely accepted that the commensal gut microbiota contributes to the health and well-being of its host. The solitary tunicate Ciona intestinalis emerges as a model organism for studying host-microbe interactions taking place in the gut, however, the potential of its gut-associated microbiota for marine biodiscovery remains unexploited. In this study, we set out to investigate the diversity, chemical space, and pharmacological potential of the gut-associated microbiota of C. intestinalis collected from the Baltic and North Seas. In a culture-based approach, we isolated 61 bacterial and 40 fungal strains affiliated to 33 different microbial genera, indicating a rich and diverse gut microbiota dominated by Gammaproteobacteria. In vitro screening of the crude microbial extracts indicated their antibacterial (64% of extracts), anticancer (22%), and/or antifungal (11%) potential. Nine microbial crude extracts were prioritized for in-depth metabolome mining by a bioactivity- and chemical diversity-based selection procedure. UPLC-MS/MS-based metabolomics combining automated (feature-based molecular networking and in silico dereplication) and manual approaches significantly improved the annotation rates. A high chemical diversity was detected where peptides and polyketides were the predominant classes. Many compounds remained unknown, including two putatively novel lipopeptides produced by a Trichoderma sp. strain. This is the first study assessing the chemical and pharmacological profile of the cultivable gut microbiota of C. intestinalis.