Structural and Evolutionary Relationships of Melanin Cascade Proteins in Cnidarian Innate Immunity.

Melanin is an essential product that plays an important role in innate immunity in a variety of organisms across the animal kingdom. Melanin synthesis is performed by many organisms using the tyrosine metabolism pathway, a general pathway that utilizes a type-three copper oxidase protein, called PO-candidates (phenoloxidase candidates). While melanin synthesis is well characterized in organisms like arthropods and humans, it is not as well understood in non-model organisms such as cnidarians. With the rising anthropomorphic climate change influence on marine ecosystems, cnidarians, specifically corals, are under an increased threat of bleaching and disease. Understanding innate immune pathways, such as melanin synthesis, is vital to gaining insights into how corals may be able to fight these threats. In this study, we use comparative bioinformatic approaches to provide a comprehensive analysis of genes involved in tyrosine-mediated melanin synthesis in cnidarians. Eighteen PO-candidates representing five phyla were studied to identify their evolutionary relationship. Cnidarian species were most similar to chordates due to domain presents in the amino acid sequences. From there, functionally conserved domains in coral proteins were identified in a coral disease dataset. Five stony corals exposed to stony coral tissue loss disease were leveraged to identify eighteen putative tyrosine metabolism genes, genes with functionally conserved domains to their Homo sapiens counterpart. To put this pathway the context of coral health, putative genes were correlated to melanin concentration from tissues of stony coral species in the disease exposure dataset. In this study, tyrosinase was identified in stony corals as correlated to melanin concentrations and likely plays a key role in immunity as a resistance trait. In addition, stony coral genes were assigned to all modules within the tyrosine metabolism pathway, indicating an evolutionary conservation of this pathway across phyla. Overall, this study provides a comprehensive analysis of the genes involved in tyrosine-mediated melanin synthesis in cnidarians.

Stony coral tissue loss disease induces transcriptional signatures of in situ degradation of dysfunctional Symbiodiniaceae.

Stony coral tissue loss disease (SCTLD), one of the most pervasive and virulent coral diseases on record, affects over 22 species of reef-building coral and is decimating reefs throughout the Caribbean. To understand how different coral species and their algal symbionts (family Symbiodiniaceae) respond to this disease, we examine the gene expression profiles of colonies of five species of coral from a SCTLD transmission experiment. The included species vary in their purported susceptibilities to SCTLD, and we use this to inform gene expression analyses of both the coral animal and their Symbiodiniaceae. We identify orthologous coral genes exhibiting lineage-specific differences in expression that correlate to disease susceptibility, as well as genes that are differentially expressed in all coral species in response to SCTLD infection. We find that SCTLD infection induces increased expression of rab7, an established marker of in situ degradation of dysfunctional Symbiodiniaceae, in all coral species accompanied by genus-level shifts in Symbiodiniaceae photosystem and metabolism gene expression. Overall, our results indicate that SCTLD infection induces symbiophagy across coral species and that the severity of disease is influenced by Symbiodiniaceae identity.

Analysis of Spatial Gene Expression at the Cellular Level in Stony Corals.

Scleractinians, or stony corals, are colonial animals that possess a high regenerative capacity and a highly diverse innate immune system. As such they present the opportunity to investigate the interconnection between regeneration and immunity in a colonial animal. Understanding the relationship between regeneration and immunity in stony corals is of further interest as it has major implications for coral reef health. One method for understanding the role of innate immunity in scleractinian regeneration is in situ hybridization using RNA probes. Here we describe a protocol for in situ hybridization in adult stony corals using a digoxigenin (DIG)-labeled RNA antisense probe which can be utilized to investigate the spatial expression of immune factors during regeneration.

Adaptive variation in homologue number within transcript families promotes expression divergence in reef-building coral.

Gene expression, especially in multispecies experiments, is used to gain insight into the genetic basis of how organisms adapt and respond to changing environments. However, evolutionary processes that can influence gene expression patterns between species such as the presence of paralogues which arise from gene duplication events are rarely accounted for. Paralogous transcripts can alter the transcriptional output of a gene, and thus exclusion of these transcripts can obscure important biological differences between species. To address this issue, we investigated how differences in transcript family size are associated with divergent gene expression patterns in five species of Caribbean reef-building corals. We demonstrate that transcript families that are rapidly evolving in terms of size have increased levels of expression divergence. Additionally, these rapidly evolving transcript families are enriched for multiple biological processes, with genes involved in the coral innate immune system demonstrating pronounced variation in homologue number between species. Overall, this investigation demonstrates the importance of incorporating paralogous transcripts when comparing gene expression across species by influencing both transcriptional output and the number of transcripts within biological processes. As this investigation was based on transcriptome assemblies, additional insights into the relationship between gene duplications and expression patterns will probably emergence once more genome assemblies are available for study.

Cnidarian Pattern Recognition Receptor Repertoires Reflect Both Phylogeny and Life History Traits.

Pattern recognition receptors (PRRs) are evolutionarily ancient and crucial components of innate immunity, recognizing danger-associated molecular patterns (DAMPs) and activating host defenses. Basal non-bilaterian animals such as cnidarians must rely solely on innate immunity to defend themselves from pathogens. By investigating cnidarian PRR repertoires we can gain insight into the evolution of innate immunity in these basal animals. Here we utilize the increasing amount of available genomic resources within Cnidaria to survey the PRR repertoires and downstream immune pathway completeness within 15 cnidarian species spanning two major cnidarian clades, Anthozoa and Medusozoa. Overall, we find that anthozoans possess prototypical PRRs, while medusozoans appear to lack these immune proteins. Additionally, anthozoans consistently had higher numbers of PRRs across all four classes relative to medusozoans, a trend largely driven by expansions in NOD-like receptors and C-type lectins. Symbiotic, sessile, and colonial cnidarians also have expanded PRR repertoires relative to their non-symbiotic, mobile, and solitary counterparts. Interestingly, cnidarians seem to lack key components of mammalian innate immune pathways, though similar to PRR numbers, anthozoans possess more complete immune pathways than medusozoans. Together, our data indicate that anthozoans have greater immune specificity than medusozoans, which we hypothesize to be due to life history traits common within Anthozoa. Overall, this investigation reveals important insights into the evolution of innate immune proteins within these basal animals.