DOI: 10.5061/dryad.sbcc2fr72
title: Parasitic cnidarians (Myxozoa) do not retain key oxygen-sensing and homeostasis tool-kit genes: Transcriptome assemblies

description: For aerobic organisms, both the Hypoxia-Inducible Factor (HIF) pathway and
 the mitochondrial genomes are key players in regulating oxygen
 homeostasis. However, recent work has suggested that these mechanisms are
 not as highly conserved as previously thought, prompting more thorough
 surveys across animal higher taxonomic levels, which would in turn permit
 testing of hypotheses about the ecological conditions that may have
 facilitated evolutionary loss of such genes. The phylum Cnidaria is known
 to harbor wide variation in mitochondrial genome morphology, from typical
 single circular chromosomes to fragmented linear chromosomes. More
 recently, members of the cnidarian clade Myxozoa, comprising obligate
 endoparasites, were shown to have lost their mitochondrial genome,
 suggesting that variation in environmental oxygen availability may be a
 key determinant in the evolution of metabolic gene networks. Here, we
 surveyed genomes and transcriptomes across 42 cnidarian species for the
 presence of HIF pathway members (HIFa, EGLN, VHL), as well as for an
 assortment of hypoxia, mitochondrial, and stress-response toolkit genes.
 We find that presence of the HIF pathway, as well as number of genes
 associated with mitochondria, hypoxia, and stress response, do not vary
 based on mitochondrial genome morphology. More interestingly, we uncover
 evidence that myxozoans have lost the canonical HIF pathway repression
 machinery, potentially altering HIF pathway functionality to work under
 the specific conditions of their parasitic lifestyles. In addition,
 relative to other cnidarians, myxozoans show loss of large proportions of
 genes associated with the mitochondrion (~39%), and involved in response
 to hypoxia (~27.5%) and general stress (~32%). Our results provide
 additional evidence that the HIF regulatory machinery is evolutionarily
 labile and that variations in the canonical system have evolved in many
 animal groups.
description: Of the 35 genome assemblies
 publicly available for Cnidaria, 17 had annotated protein sequence files
 that were used in this study. These included classes Anthozoa (<em>n
 </em>= 9), Cubozoa (1), Hydrozoa (2), Myxozoa (3), and Scyphozoa (2)
 and were downloaded directly from their respective repositories
 (Supplemental Table 1; available online). To increase taxon sampling,
 transcriptomic data (Illumina paired-end mRNA reads) from an additional 25
 species were downloaded from the NCBI Sequence Read Archive (SRA) and
 assembled <em>de novo</em>. Among these were species in the
 classes Cubozoa (4), Hydrozoa (9), Myxozoa (5), and Scyphozoa (4) and
 Staurozoa (3) (Supplemental Table 2; available online). The singular
 species of the group Polylpodium was also included. For de novo transcriptome
 assemblies, downloaded reads were trimmed for base quality (&gt;Q30)
 and adapters using BBDuk (Bushnell<em> et al.</em>, 2017), and
 transcripts assembled <em>de novo</em> with Trinity
 (Haas<em> et al.</em>, 2013) using default parameters. The
 assembled transcripts were then translated to protein sequences using
 Transdecoder, which included steps for predicting all open reading frames
 (ORFs), BLASTing of ORFs against the Uniprot/Swissprot database (release
 date March 2021), and retaining the best protein models for each
 transcript. For transcriptome assemblies from the endoparasitic myxozoan
 species, contamination from host tissue (fish or annelid) is likely to be
 present in the pool of reads, as observed for mRNA-based assemblies from
 parasitic copepods (Graham and Barreto, 2020). We hence filtered the
 myxozoan assemblies by identifying transcripts assembled from potential
 host RNA. We prepared a custom BLAST database containing protein sequences
 from three fish and one annelid obtained from Ensembl. These were
 <em>Oncorhynchus mykiss </em>(assembly Omyk_1.0, accession
 GCA_002163495.1), <em>Cyprinus carpio</em>
 (common_carp_genome, GCA_000951615.2), <em>Sparus aurata
 </em>(SpaAur1.1, GCA_900880675.1), and <em>Capitella
 teleta</em> (Capitella telata v1.0, GCA_000328365.1). We then
 performed BLASTP searches of our assembled myxozoan sequences against this
 database. Hits that had a match within these potential host sequences,
 with <em>e-</em>value ≤ 1e-75 and percent identity ≥ 85, were
 considered contaminant and removed from the assemblies, as used by
 Yahalomi<em> et al.</em> (2020).
description: Includes all transcriptomes used in this study - both de novo
 generated and from other sources.