Evolution of Methyltransferase-Like (METTL) Proteins in Metazoa: A Complex Gene Family Involved in Epitranscriptomic Regulation and Other Epigenetic Processes

The methyltransferase-like (METTL) proteins constitute a family of seven-beta-strand methyltransferases with S-adenosyl methionine-binding domains that modify DNA, RNA, and proteins. Methylation by METTL proteins contributes to the epigenetic, and in the case of RNA modifications, epitranscriptomic regulation of a variety of biological processes. Despite their functional importance, most investigations of the substrates and functions of METTLs within metazoans have been restricted to model vertebrate taxa. In the present work, we explore the evolutionary mechanisms driving the diversification and functional differentiation of 33 individual METTL proteins across Metazoa. Our results show that METTLs are nearly ubiquitous across the animal kingdom, with most having arisen early in metazoan evolution (i.e., occur in basal metazoan phyla). Individual METTL lineages each originated from single independent ancestors, constituting monophyletic clades, which suggests that each METTL was subject to strong selective constraints driving its structural and/or functional specialization. Interestingly, a similar process did not extend to the differentiation of nucleoside-modifying and protein-modifying METTLs (i.e., each METTL type did not form a unique monophyletic clade). The members of these two types of METTLs also exhibited differences in their rates of evolution. Overall, we provide evidence that the long-term evolution of METTL family members was driven by strong purifying selection, which in combination with adaptive selection episodes, led to the functional specialization of individual METTL lineages. This work contributes useful information regarding the evolution of a gene family that fulfills a variety of epigenetic functions, and can have profound influences on molecular processes and phenotypic traits.

Molecular mechanisms of sperm motility are conserved in a basal metazoan

Efficient and targeted sperm motility is essential for animal reproductive success. Studies in mammals and echinoderms have uncovered a highly conserved signaling mechanism in which sperm motility is stimulated by pH-dependent activation of the cAMP-producing enzyme soluble adenylyl cyclase (sAC). However, the presence of this pathway in basal metazoans has, until now, been unexplored. Here we found that cytoplasmic alkalinization induced a rapid burst of cAMP signaling and the full activation of motility in sperm from the reef-building coral Montipora capitata. Coral sperm expressed sAC in the flagellum, midpiece, and acrosomal regions, indicating that this molecular pH sensor may play a role in regulating mitochondrial respiration and flagellar beating. In bilaterians, sAC is a central node of a broader pH-dependent signaling pathway that alters cellular behavior in response to changes to the extracellular environment. We present transcript-level evidence that a homologous pathway is present in coral sperm, including the Na+/H+ exchanger SLC9C1, protein kinase A, and the CatSper Ca2+ channel conserved even in mammalian sperm. Our discovery of this pathway in a basal metazoan species highlights the ancient origin of the pH-sAC-cAMP signaling node in sperm physiology and suggests that it may be present in many other marine invertebrate taxa for which sperm motility mechanisms remain unexplored. These results emphasize our need to better understand the role of pH-dependent signaling in marine reproductive success, particularly as worsening ocean acidification and warming due to climate change continue to impair the physiology of corals and other marine invertebrates.

Evolutionary Analysis of Cystatins of Early-Emerging Metazoans Reveals a Novel Subtype in Parasitic Cnidarians

The evolutionary aspects of cystatins are greatly underexplored in early-emerging metazoans. Thus, we surveyed the gene organization, protein architecture, and phylogeny of cystatin homologues mined from 110 genomes and the transcriptomes of 58 basal metazoan species, encompassing free-living and parasite taxa of Porifera, Placozoa, Cnidaria (including Myxozoa), and Ctenophora. We found that the cystatin gene repertoire significantly differs among phyla, with stefins present in most of the investigated lineages but with type 2 cystatins missing in several basal metazoan groups. Similar to liver and intestinal flukes, myxozoan parasites possess atypical stefins with chimeric structure that combine motifs of classical stefins and type 2 cystatins. Other early metazoan taxa regardless of lifestyle have only the classical representation of cystatins and lack multi-domain ones. Our comprehensive phylogenetic analyses revealed that stefins and type 2 cystatins clustered into taxonomically defined clades with multiple independent paralogous groups, which probably arose due to gene duplications. The stefin clade split between the subclades of classical stefins and the atypical stefins of myxozoans and flukes. Atypical stefins represent key evolutionary innovations of the two parasite groups for which their origin might have been linked with ancestral gene chimerization, obligate parasitism, life cycle complexity, genome reduction, and host immunity.

Spontaneous electrical low-frequency oscillations: a possible role in Hydra and all living systems

As one of the first model systems in biology, the basal metazoan Hydra has been revealing fundamental features of living systems since it was first discovered by Antonie van Leeuwenhoek in the early eighteenth century. While it has become well-established within cell and developmental biology, this tiny freshwater polyp is only now being re-introduced to modern neuroscience where it has already produced a curious finding: the presence of low-frequency spontaneous neural oscillations at the same frequency as those found in the default mode network in the human brain. Surprisingly, increasing evidence suggests such spontaneous electrical low-frequency oscillations (SELFOs) are found across the wide diversity of life on Earth, from bacteria to humans. This paper reviews the evidence for SELFOs in diverse phyla, beginning with the importance of their discovery in Hydra , and hypothesizes a potential role as electrical organism organizers, which supports a growing literature on the role of bioelectricity as a ‘template’ for developmental memory in organism regeneration. This article is part of the theme issue ‘Basal cognition: conceptual tools and the view from the single cell’.

Ancestral role of TNF-R pathway in cell differentiation in the basal metazoan Hydra

Tumour necrosis factor receptors (TNF-Rs) and their ligands, tumour necrosis factors are highly conserved proteins described in all metazoan phyla. They function as inducers of extrinsic apoptotic signalling and facilitate inflammation, differentiation and cell survival. TNF-Rs use distinct adaptor molecules to activate signalling cascades. FADD-adaptors often mediate apoptosis and TRAF-adaptors mediate cell differentiation and inflammation. Most of these pathway components are conserved in cnidarians and here we investigated the Hydra TNF-R. We report that it is related to the ectodysplasin receptor, which is involved in epithelial cell differentiation in mammals. In Hydra, it is localised in epithelial cells with incorporated nematocytes in tentacles and body column, indicating a similar function. Further experiments suggest that it interacts with the Hydra homolog of a TRAF-adaptor, but not with FADDs. Hydra-FADD proteins co-localised with Hydra caspases in death effector filaments and recruited caspases suggesting that they are part of an apoptotic signalling pathway. Regulating epithelial cell differentiation via TRAF-adaptors therefore seems to be an ancient function of TNF-Rs, whereas FADD-caspase interactions may be part of a separate apoptotic pathway.

The Native Microbiome is Crucial for Offspring Generation and Fitness of Aurelia Aurita

ABSTRACT All multicellular organisms are associated with microbial communities, ultimately forming a metaorganism. Several studies conducted on well-established model organisms point to immunological, metabolic, and behavioral benefits of the associated microbiota for the host. Consequently, a microbiome can influence the physiology of a host; moreover, microbial community shifts can affect host health and fitness. The present study aimed to evaluate the significance and functional role of the native microbiota for life cycle transitions and fitness of the cnidarian moon jellyfish Aurelia aurita. A comprehensive host fitness experiment was conducted studying the polyp life stage and integrating 12 combinations of treatments with microbiota modification (sterile conditions, foreign food bacteria, and potential pathogens). Asexual reproduction, e.g., generation of daughter polyps, and the formation and release of ephyrae were highly affected in the absence of the native microbiota, ultimately resulting in a halt of strobilation and ephyra release. Assessment of further fitness traits showed that health, growth, and feeding rate were decreased in the absence and upon community changes of the native microbiota, e.g., when challenged with selected bacteria. Moreover, changes in microbial community patterns were detected by 16S rRNA amplicon sequencing during the course of the experiment. This demonstrated that six operational taxonomic units (OTUs) significantly correlated and explained up to 97% of fitness data variability, strongly supporting the association of impaired fitness with the absence/presence of specific bacteria. Conclusively, our study provides new insights into the importance and function of the microbiome for asexual reproduction, health, and fitness of the basal metazoan A. aurita. IMPORTANCE All multicellular organisms are associated with a diverse and specific community of microorganisms; consequently, the microbiome is of fundamental importance for health and fitness of the multicellular host. However, studies on microbiome contribution to host fitness are in their infancy, in particular, for less well-established hosts such as the moon jellyfish Aurelia aurita. Here, we studied the impact of the native microbiome on the asexual reproduction and on further fitness traits (health, growth, and feeding) of the basal metazoan due to induced changes in its microbiome. We observed significant impact on all fitness traits analyzed, in particular, in the absence of the protective microbial shield and when challenged with marine potentially pathogenic bacterial isolates. Notable is the identified crucial importance of the native microbiome for the generation of offspring, consequently affecting life cycle decisions. Thus, we conclude that the microbiome is essential for the maintenance of a healthy metaorganism.

Differential expression of Exaiptasia pallida GIMAP genes upon induction of apoptosis and autophagy suggests a potential role in cnidarian symbiosis and disease

ABSTRACTCoral reefs, one of the world's most productive and diverse ecosystems, are currently threatened by a variety of stressors that result in increased prevalence of both bleaching and disease. Therefore, understanding the molecular mechanisms involved in these responses is critical to mitigate future damage to the reefs. One group of genes that is potentially involved in cnidarian immunity and symbiosis is GTPases of immunity associated proteins (GIMAP). In vertebrates, this family of proteins is involved in regulating the fate of developing lymphocytes and interacts with proteins involved in apoptosis and autophagy. As apoptosis, autophagy and immunity have previously been shown to be involved in cnidarian symbiosis and disease, the goal of this research was to determine the role of cnidarian GIMAPs in these processes using the anemone Exaiptasia pallida. To do so, GIMAP genes were characterized in the E. pallida genome and changes in gene expression were measured using qPCR in response to chemical induction of apoptosis, autophagy and treatment with the immune stimulant lipopolysaccharide (LPS) in both aposymbiotic and symbiotic anemones. The results revealed four GIMAP-like genes in E. pallida, referred to as Ep_GIMAPs. Induction of apoptosis and autophagy resulted in a general downregulation of Ep_GIMAPs, but no significant changes were observed in response to LPS treatment. This indicates that Ep_GIMAPs may be involved in the regulation of apoptosis and autophagy, and therefore could play a role in cnidarian–dinoflagellate symbiosis. Overall, these results increase our knowledge on the function of GIMAPs in a basal metazoan.

Monophyletic Origin and Divergent Evolution of Animal Telomerase RNA

Telomerase RNA (TR) is a noncoding RNA essential for the function of telomerase ribonucleoprotein. TRs from vertebrates, fungi, ciliates, and plants exhibit extreme diversity in size, sequence, secondary structure, and biogenesis pathway. However, the evolutionary pathways leading to such unusual diversity among eukaryotic kingdoms remain elusive. Within the metazoan kingdom, the study of TR has been limited to vertebrates and echinoderms. To understand the origin and evolution of TR across the animal kingdom, we employed a phylogeny-guided, structure-based bioinformatics approach to identify 82 novel TRs from eight previously unexplored metazoan phyla, including the basal-branching sponges. Synthetic TRs from two representative species, a hemichordate and a mollusk, reconstitute active telomerase in vitro with their corresponding telomerase reverse transcriptase components, confirming that they are authentic TRs. Comparative analysis shows that three functional domains, template-pseudoknot (T-PK), CR4/5, and box H/ACA, are conserved between vertebrate and the basal metazoan lineages, indicating a monophyletic origin of the animal TRs with a snoRNA-related biogenesis mechanism. Nonetheless, TRs along separate animal lineages evolved with divergent structural elements in the T-PK and CR4/5 domains. For example, TRs from echinoderms and protostomes lack the canonical CR4/5 and have independently evolved functionally equivalent domains with different secondary structures. In the T-PK domain, a P1.1 stem common in most metazoan clades defines the template boundary, which is replaced by a P1-defined boundary in vertebrates. This study provides unprecedented insight into the divergent evolution of detailed TR secondary structures across broad metazoan lineages, revealing ancestral and later-diversified elements.

Prototypical pacemaker neurons interact with the resident microbiota

Pacemaker neurons exert control over neuronal circuit function by their intrinsic ability to generate rhythmic bursts of action potential. Recent work has identified rhythmic gut contractions in human, mice, and hydra to be dependent on both neurons and the resident microbiota. However, little is known about the evolutionary origin of these neurons and their interaction with microbes. In this study, we identified and functionally characterized prototypical ANO/SCN/TRPM ion channel-expressing pacemaker cells in the basal metazoanHydraby using a combination of single-cell transcriptomics, immunochemistry, and functional experiments. Unexpectedly, these prototypical pacemaker neurons express a rich set of immune-related genes mediating their interaction with the microbial environment. Furthermore, functional experiments gave a strong support to a model of the evolutionary emergence of pacemaker cells as neurons using components of innate immunity to interact with the microbial environment and ion channels to generate rhythmic contractions.