Functional analyses of bacterial genomes found in Symbiodiniaceae genome assemblies.

Bacterial-algal interactions strongly influence marine ecosystems. Bacterial communities in cultured dinoflagellates of the family Symbiodiniaceae have been characterized by metagenomics. However, little is known about whole-genome analysis of marine bacteria associated with these dinoflagellates. We performed in silico analysis of four bacterial genomes from cultures of four dinoflagellates of the genera Symbiodinium, Breviolum, Cladocopium and Durusdinium. Comparative analysis showed that the former three contain the alphaproteobacterial family Parvibaculaceae and that the Durusdinium culture includes the family Sphingomonadaceae. There were no large genomic reductions in the alphaproteobacteria with genome sizes of 2.9-3.9 Mb, implying they are not obligate intracellular bacteria. Genomic annotations of three Parvibaculaceae detected the gene for diacetylchitobiose deacetylase (Dac), which may be involved in the degradation of dinoflagellate cell surfaces. They also had metabolic genes for dissimilatory nitrate reduction to ammonium (DNRA) in the nitrogen (N) cycle and cobalamin (vitamin B12 ) biosynthetic genes in the salvage pathway. Those three characters were not found in the Sphingomonadaceae genome. Predicted biosynthetic gene clusters for secondary metabolites indicated that the Parvibaculaceae likely produce the same secondary metabolites. Our study suggests that the Parvibaculaceae is a major resident of Symbiodiniaceae cultures with antibiotics.

In Vitro Phagocytosis of Different Dinoflagellate Species by Coral Cells.

Coral-dinoflagellate symbiosis is a unique biological phenomenon, in which animal cells engulf single-celled photosynthetic algae and maintain them in their cytoplasm mutualistically. Studies are needed to reveal the complex mechanisms involved in symbiotic processes, but it is difficult to answer these questions using intact corals. To tackle these issues, our previous studies established an in vitro system of symbiosis between cells of the scleractinian coral Acropora tenuis and the dinoflagellate Breviolum minutum, and showed that corals direct phagocytosis, while algae are likely engulfed by coral cells passively. Several genera of the family Symbiodiniaceae can establish symbioses with corals, but the symbiotic ratio differs depending on the dinoflagellate clades involved. To understand possible causes of these differences, this study examined whether cultured coral cells show phagocytotic activity with various dinoflagellate strains similar to those shown by intact A. tenuis. We found that (a) A. tenuis larvae incorporate Symbiodinium and Breviolum, but not Cladocopium, and very few Effrenium, (b) cultured coral cells engulfed all four species but the ratio of engulfment was significantly higher with Symbiodinium and Breviolum than Cladocopium and Effrenium, (c) cultured coral cells also phagocytosed inorganic latex beads differently than they do dinoflagellates . It is likely that cultured coral cells preferentially phagocytose Symbiodinium and Breviolum, suggesting that specific molecular mechanisms involved in initiation of symbiosis should be investigated in the future.

Building consensus around the assessment and interpretation of Symbiodiniaceae diversity.

Within microeukaryotes, genetic variation and functional variation sometimes accumulate more quickly than morphological differences. To understand the evolutionary history and ecology of such lineages, it is key to examine diversity at multiple levels of organization. In the dinoflagellate family Symbiodiniaceae, which can form endosymbioses with cnidarians (e.g., corals, octocorals, sea anemones, jellyfish), other marine invertebrates (e.g., sponges, molluscs, flatworms), and protists (e.g., foraminifera), molecular data have been used extensively over the past three decades to describe phenotypes and to make evolutionary and ecological inferences. Despite advances in Symbiodiniaceae genomics, a lack of consensus among researchers with respect to interpreting genetic data has slowed progress in the field and acted as a barrier to reconciling observations. Here, we identify key challenges regarding the assessment and interpretation of Symbiodiniaceae genetic diversity across three levels: species, populations, and communities. We summarize areas of agreement and highlight techniques and approaches that are broadly accepted. In areas where debate remains, we identify unresolved issues and discuss technologies and approaches that can help to fill knowledge gaps related to genetic and phenotypic diversity. We also discuss ways to stimulate progress, in particular by fostering a more inclusive and collaborative research community. We hope that this perspective will inspire and accelerate coral reef science by serving as a resource to those designing experiments, publishing research, and applying for funding related to Symbiodiniaceae and their symbiotic partnerships.

Active Expression of Genes for Protein Modification Enzymes in Habu Venom Glands.

Genes encoding snake venom toxins have been studied extensively. However, genes involved in the modification and functioning of venom proteins are little known. Protobothrops is a genus of pit vipers, which are venomous and inhabit the Nansei (Southwest) islands of Japan, Taiwan China, Vietnam, Thailand, Myanmar, Nepal, Bhutan, and India. Our previous study decoded the genome of Protobothrops flavoviridis, a species endemic to the Nansei Islands, Japan, and revealed unique evolutionary processes of some venom genes. In this study, we analyzed genes that are highly expressed in venom glands to survey genes for candidate enzymes or chaperone proteins involved in toxin folding and modification. We found that, in addition to genes that encode venom proteins and ribosomal proteins, genes that encode protein disulfide isomerase (PDI) family members (orthologs of human P4HB and PDIA3), Selenoprotein M (SELENOM), and Calreticulin (CALR) are highly expressed in venom glands. Since these enzymes or chaperones are involved in protein modification and potentially possess protein folding functions, we propose that P4HB, SELENOM, CALR, and PDIA3 encode candidate enzymes or chaperones to confer toxic functions upon the venom transcriptome.

Gene clusters for biosynthesis of mycosporine-like amino acids in dinoflagellate nuclear genomes: Possible recent horizontal gene transfer between species of Symbiodiniaceae (Dinophyceae).

Global warming increases the temperature of the ocean surface, which can disrupt dinoflagellate-coral symbioses and result in coral bleaching. Photosynthetic dinoflagellates of the family Symbiodiniaceae include bleaching-tolerant and bleaching-sensitive coral symbionts. Therefore, understanding the molecular mechanisms for changing symbiont diversity is potentially useful to assist recovery of coral holobionts (corals and their associated microbes, including multiple species of Symbiodiniaceae), although sexual reproduction has not been observed in the Symbiodiniaceae. Recent molecular phylogenetic analyses estimate that the Symbiodiniaceae appeared 160 million years ago and diversified into 15 groups, five genera of which now have available draft genomes (i.e., Symbiodinium, Durusdinium, Breviolum, Fugacium, and Cladocopium). Comparative genomic analyses have suggested that crown groups have fewer gene families than early-diverging groups, although many genes that were probably acquired via gene duplications and horizontal gene transfers (HGTs) have been found in each decoded genome. Because UV stress is likely a contributor to coral bleaching, and because the highly conserved gene cluster for mycosporine-like amino acid (MAA) biosynthesis has been found in thermal-tolerant symbiont genomes, I reviewed genomic features of the Symbiodiniaceae, focusing on possible acquisition of a biosynthetic gene cluster for MAAs, which absorb UV radiation. On the basis of highly conserved noncoding sequences, I hypothesized that HGTs have occurred among members of the Symbiodiniaceae and have contributed to the diversification of Symbiodiniaceae-host relationships. Finally, I proposed that bleaching tolerance may be strengthened by multiple MAAs from both symbiotic dinoflagellates and corals.

Coral symbionts evolved a functional polycistronic flavodiiron gene.

Photosynthesis in cyanobacteria, green algae, and basal land plants is protected against excess reducing pressure on the photosynthetic chain by flavodiiron proteins (FLV) that dissipate photosynthetic electrons by reducing O2. In these organisms, the genes encoding FLV are always conserved in the form of a pair of two-type isozymes (FLVA and FLVB) that are believed to function in O2 photo-reduction as a heterodimer. While coral symbionts (dinoflagellates of the family Symbiodiniaceae) are the only algae to harbor FLV in photosynthetic red plastid lineage, only one gene is found in transcriptomes and its role and activity remain unknown. Here, we characterized the FLV genes in Symbiodiniaceae and found that its coding region is composed of tandemly repeated FLV sequences. By measuring the O2-dependent electron flow and P700 oxidation, we suggest that this atypical FLV is active in vivo. Based on the amino-acid sequence alignment and the phylogenetic analysis, we conclude that in coral symbionts, the gene pair for FLVA and FLVB have been fused to construct one coding region for a hybrid enzyme, which presumably occurred when or after both genes were inherited from basal green algae to the dinoflagellate. Immunodetection suggested the FLV polypeptide to be cleaved by a post-translational mechanism, adding it to the rare cases of polycistronic genes in eukaryotes. Our results demonstrate that FLV are active in coral symbionts with genomic arrangement that is unique to these species. The implication of these unique features on their symbiotic living environment is discussed.

Genome and transcriptome assemblies of the kuruma shrimp, Marsupenaeus japonicus.

The kuruma shrimp Marsupenaeus japonicus (order Decapoda, family Penaeidae) is an economically important crustacean that occurs in shallow, warm seas across the Indo-Pacific. Here, using a combination of Illumina and Oxford Nanopore Technologies platforms, we produced a draft genome assembly of M. japonicus (1.70 Gbp; 18,210 scaffolds; scaffold N50 = 234.9 kbp; 34.38% GC, 93.4% BUSCO completeness) and a complete mitochondrial genome sequence (15,969 bp). As with other penaeid shrimp genomes, the M. japonicus genome is extremely rich in simple repeats, which occupies 27.4% of the assembly. A total of 26,381 protein-coding gene models (94.7% BUSCO completeness) were predicted, of which 18,005 genes (68.2%) were assigned functional description by at least one method. We also produced an Illumina-based transcriptome shotgun assembly (40,991 entries; 93.0% BUSCO completeness) and a PacBio Iso-Seq transcriptome assembly (25,415 entries; 67.5% BUSCO completeness). We envision that the M. japonicus genome and transcriptome assemblies will serve as useful resources for the basic research, fisheries management, and breeding programs of M. japonicus.

Expansion and Diversification of Fluorescent Protein Genes in Fifteen Acropora Species during the Evolution of Acroporid Corals.

In addition to a purple, non-fluorescent chromoprotein (ChrP), fluorescent proteins (FPs) account for the vivid colors of corals, which occur in green (GFP), cyan (CFP), and red (RFP) FPs. To understand the evolution of the coral FP gene family, we examined the genomes of 15 Acropora species and three confamilial taxa. This genome-wide survey identified 219 FP genes. Molecular phylogeny revealed that the 15 Acropora species each have 9-18 FP genes, whereas the other acroporids examined have only two, suggesting a pronounced expansion of the FP genes in the genus Acropora. The data estimates of FP gene duplication suggest that the last common ancestor of the Acropora species that survived in the period of high sea surface temperature (Paleogene period) has already gained 16 FP genes. Different evolutionary histories of lineage-specific duplication and loss were discovered among GFP/CFPs, RFPs, and ChrPs. Synteny analysis revealed core GFP/CFP, RFP, and ChrP gene clusters, in which a tandem duplication of the FP genes was evident. The expansion and diversification of Acropora FPs may have contributed to the present-day richness of this genus.

Establishing Sustainable Cell Lines of a Coral, Acropora tenuis.

Planula larvae of the scleractinian coral, Acropora tenuis, consist of elongated ectodermal cells and developing inner endodermal cells. To establish in vitro cell lines for future studies of cellular and developmental potential of coral cells, larvae were successfully dissociated into single cells by treating them with a tissue dissociation solution consisting of trypsin, EDTA, and collagenase. Brown-colored cells, translucent cells, and pale blue cells were the major components of dissociated larvae. Brown-colored cells began to proliferate transiently in the culture medium that was devised for the coral, while translucent cells and pale blue cells decreased in number about 1 week after cell dissociation. In addition, when a modular protease, plasmin, was added to the cell culture medium, brown-colored cells extended pseudopodia and assumed amorphous shapes. They then continued to proliferate in clumps for more than 6 months with a doubling time of approximately 4-5 days. From 3 weeks of cell culture onward, brown-colored cells often aggregated and exhibited morphogenesis-like behavior to form flat sheets, and blastula-like clusters or gastrula-like spheres. Single cells or cell-clusters of the cell lines were analyzed by RNA-seq. This analysis showed that genes expressed in these cells in vitro were A. tenuis genes. Furthermore, each cell line expressed a specific set of genes, suggesting that their properties include gastroderm, secretory cells, undifferentiated cells, neuronal cells, and epidermis. All cell properties were maintained stably throughout successive cell cultures. These results confirm the successful establishment of a coral in vitro cell line.

Application of omics research in seaweeds with a focus on red seaweeds.

Targeted 'omics' research for seaweeds, utilizing various computational and informatics frameworks, has the potential to rapidly develop our understanding of biological processes at the molecular level and contribute to solutions for the most pressing environmental and social issues of our time. Here, a systematic review into the current status of seaweed omics research was undertaken to evaluate the biological diversity of seaweed species investigated (red, green and brown phyla), the levels to which the work was undertaken (from full genome to transcripts, proteins or metabolites) and the field of research to which it has contributed. We report that from 1994 to 2021 the majority of seaweed omics research has been performed on the red seaweeds (45% of total studies), with more than half of these studies based upon two genera Pyropia and Gracilaria. A smaller number of studies examined brown seaweed (key genera Saccharina and Sargassum) and green seaweed (primarily Ulva). Overall, seaweed omics research is most highly associated with the field of evolution (46% of total studies), followed by the fields of ecology, natural products and their biosynthesis, omics methodology and seaweed-microbe interactions. Synthesis and specific outcomes derived from omics studies in the red seaweeds are provided. Together, these studies have provided a broad-scale interrogation of seaweeds, facilitating our ability to answer fundamental queries and develop applied outcomes. Crucial to the next steps will be establishing analytical tools and databases that can be more broadly utilized by practitioners and researchers across the globe because of their shared interest in the key seaweed genera.

Whole-Genome Transcriptome Analyses of Native Symbionts Reveal Host Coral Genomic Novelties for Establishing Coral-Algae Symbioses.

Reef-building corals and photosynthetic, endosymbiotic algae of the family Symbiodiniaceae establish mutualistic relationships that are fundamental to coral biology, enabling coral reefs to support a vast diversity of marine species. Although numerous types of Symbiodiniaceae occur in coral reef environments, Acropora corals select specific types in early life stages. In order to study molecular mechanisms of coral-algal symbioses occurring in nature, we performed whole-genome transcriptomic analyses of Acropora tenuis larvae inoculated with Symbiodinium microadriaticum strains isolated from an Acropora recruit. In order to identify genes specifically involved in symbioses with native symbionts in early life stages, we also investigated transcriptomic responses of Acropora larvae exposed to closely related, nonsymbiotic, and occasionally symbiotic Symbiodinium strains. We found that the number of differentially expressed genes was largest when larvae acquired native symbionts. Repertoires of differentially expressed genes indicated that corals reduced amino acid, sugar, and lipid metabolism, such that metabolic enzymes performing these functions were derived primarily from S. microadriaticum rather than from A. tenuis. Upregulated gene expression of transporters for those metabolites occurred only when coral larvae acquired their natural symbionts, suggesting active utilization of native symbionts by host corals. We also discovered that in Acropora, genes for sugar and amino acid transporters, prosaposin-like, and Notch ligand-like, were upregulated only in response to native symbionts, and included tandemly duplicated genes. Gene duplications in coral genomes may have been essential to establish genomic novelties for coral-algae symbiosis.

A New Dinoflagellate Genome Illuminates a Conserved Gene Cluster Involved in Sunscreen Biosynthesis.

Photosynthetic dinoflagellates of the Family Symbiodiniaceae live symbiotically with many organisms that inhabit coral reefs and are currently classified into fifteen groups, including seven genera. Draft genomes from four genera, Symbiodinium, Breviolum, Fugacium, and Cladocopium, which have been isolated from corals, have been reported. However, no genome is available from the genus Durusdinium, which occupies an intermediate phylogenetic position in the Family Symbiodiniaceae and is well known for thermal tolerance (resistance to bleaching). We sequenced, assembled, and annotated the genome of Durusdinium trenchii, isolated from the coral, Favia speciosa, in Okinawa, Japan. Assembled short reads amounted to 670 Mb with ∼47% GC content. This GC content was intermediate among taxa belonging to the Symbiodiniaceae. Approximately 30,000 protein-coding genes were predicted in the D. trenchii genome, fewer than in other genomes from the Symbiodiniaceae. However, annotations revealed that the D. trenchii genome encodes a cluster of genes for synthesis of mycosporine-like amino acids, which absorb UV radiation. Interestingly, a neighboring gene in the cluster encodes a glucose-methanol-choline oxidoreductase with a flavin adenine dinucleotide domain that is also found in Symbiodinium tridacnidorum. This conservation seems to partially clarify an ancestral genomic structure in the Symbiodiniaceae and its loss in late-branching lineages, including Breviolum and Cladocopium, after splitting from the Durusdinium lineage. Our analysis suggests that approximately half of the taxa in the Symbiodiniaceae may maintain the ability to synthesize mycosporine-like amino acids. Thus, this work provides a significant genomic resource for understanding the genomic diversity of Symbiodiniaceae in corals.

Response of Coral Reef Dinoflagellates to Nanoplastics under Experimental Conditions Suggests Downregulation of Cellular Metabolism.

Plastic products contribute heavily to anthropogenic pollution of the oceans. Small plastic particles in the microscale and nanoscale ranges have been found in all marine ecosystems, but little is known about their effects upon marine organisms. In this study, we examine changes in cell growth, aggregation, and gene expression of two symbiotic dinoflagellates of the family Symbiodiniaceae, Symbiodinium tridacnidorum (clade A3), and Cladocopium sp. (clade C) under exposure to 42-nm polystyrene beads. In laboratory experiments, the cell number and aggregation were reduced after 10 days of nanoplastic exposure at 0.01, 0.1, and 10 mg/L concentrations, but no clear correlation with plastic concentration was observed. Genes involved in dynein motor function were upregulated when compared to control conditions, while genes related to photosynthesis, mitosis, and intracellular degradation were downregulated. Overall, nanoplastic exposure led to more genes being downregulated than upregulated and the number of genes with altered expression was larger in Cladocopium sp. than in S. tridacnidorum, suggesting different sensitivity to nano-plastics between species. Our data show that nano-plastic inhibits growth and alters aggregation properties of microalgae, which may negatively affect the uptake of these indispensable symbionts by coral reef organisms.

Integrated omics unveil the secondary metabolic landscape of a basal dinoflagellate.

BACKGROUND: Some dinoflagellates cause harmful algal blooms, releasing toxic secondary metabolites, to the detriment of marine ecosystems and human health. Our understanding of dinoflagellate toxin biosynthesis has been hampered by their unusually large genomes. To overcome this challenge, for the first time, we sequenced the genome, microRNAs, and mRNA isoforms of a basal dinoflagellate, Amphidinium gibbosum, and employed an integrated omics approach to understand its secondary metabolite biosynthesis. RESULTS: We assembled the ~ 6.4-Gb A. gibbosum genome, and by probing decoded dinoflagellate genomes and transcriptomes, we identified the non-ribosomal peptide synthetase adenylation domain as essential for generation of specialized metabolites. Upon starving the cells of phosphate and nitrogen, we observed pronounced shifts in metabolite biosynthesis, suggestive of post-transcriptional regulation by microRNAs. Using Iso-Seq and RNA-seq data, we found that alternative splicing and polycistronic expression generate different transcripts for secondary metabolism. CONCLUSIONS: Our genomic findings suggest intricate integration of various metabolic enzymes that function iteratively to synthesize metabolites, providing mechanistic insights into how dinoflagellates synthesize secondary metabolites, depending upon nutrient availability. This study provides insights into toxin production associated with dinoflagellate blooms. The genome of this basal dinoflagellate provides important clues about dinoflagellate evolution and overcomes the large genome size, which has been a challenge previously.

Comparative genomics of four strains of the edible brown alga, Cladosiphon okamuranus.

BACKGROUND: The brown alga, Cladosiphon okamuranus (Okinawa mozuku), is one of the most important edible seaweeds, and it is cultivated for market primarily in Okinawa, Japan. Four strains, denominated S, K, O, and C, with distinctively different morphologies, have been cultivated commercially since the early 2000s. We previously reported a draft genome of the S-strain. To facilitate studies of seaweed biology for future aquaculture, we here decoded and analyzed genomes of the other three strains (K, O, and C). RESULTS: Here we improved the genome of the S-strain (ver. 2, 130 Mbp, 12,999 genes), and decoded the K-strain (135 Mbp, 12,511 genes), the O-strain (140 Mbp, 12,548 genes), and the C-strain (143 Mbp, 12,182 genes). Molecular phylogenies, using mitochondrial and nuclear genes, showed that the S-strain diverged first, followed by the K-strain, and most recently the C- and O-strains. Comparisons of genome architecture among the four strains document the frequent occurrence of inversions. In addition to gene acquisitions and losses, the S-, K-, O-, and C-strains possess 457, 344, 367, and 262 gene families unique to each strain, respectively. Comprehensive Blast searches showed that most genes have no sequence similarity to any entries in the non-redundant protein sequence database, although GO annotation suggested that they likely function in relation to molecular and biological processes and cellular components. CONCLUSIONS: Our study compares the genomes of four strains of C. okamuranus and examines their phylogenetic relationships. Due to global environmental changes, including temperature increases, acidification, and pollution, brown algal aquaculture is facing critical challenges. Genomic and phylogenetic information reported by the present research provides useful tools for isolation of novel strains.

Correlation between Organelle Genetic Variation and RNA Editing in Dinoflagellates Associated with the Coral Acropora digitifera.

In order to develop successful strategies for coral reef preservation, it is critical that the biology of both host corals and symbiotic algae are investigated. In the Ryukyu Archipelago, which encompasses many islands spread over ∼500 km of the Pacific Ocean, four major populations of the coral Acropora digitifera have been studied using whole-genome shotgun (WGS) sequence analysis (Shinzato C, Mungpakdee S, Arakaki N, Satoh N. 2015. Genome-wide single-nucleotide polymorphism (SNP) analysis explains coral diversity and recovery in the Ryukyu Archipelago. Sci Rep. 5:18211.). In contrast, the diversity of the symbiotic dinoflagellates associated with these A. digitifera populations is unknown. It is therefore unclear if these two core components of the coral holobiont share a common evolutionary history. This issue can be addressed for the symbiotic algal populations by studying the organelle genomes of their mitochondria and plastids. Here, we analyzed WGS data from ∼150 adult A. digitifera, and by mapping reads to the available reference genome sequences, we extracted 2,250 sequences representing 15 organelle genes of Symbiodiniaceae. Molecular phylogenetic analyses of these mitochondrial and plastid gene sets revealed that A. digitifera from the southern Yaeyama islands harbor a different Symbiodiniaceae population than the islands of Okinawa and Kerama in the north, indicating that the distribution of symbiont populations partially matches that of the four host populations. Interestingly, we found that numerous SNPs correspond to known RNA-edited sites in 14 of the Symbiodiniaceae organelle genes, with mitochondrial genes showing a stronger correspondence than plastid genes. These results suggest a possible correlation between RNA editing and SNPs in the two organelle genomes of symbiotic dinoflagellates.

Organelle inheritance and genome architecture variation in isogamous brown algae.

Among the brown algal lineages, Ectocarpales species have isogamous fertilization in which male and female gametes are morphologically similar. In contrast, female gametes are much larger than male gametes in the oogamous species found in many other brown algal lineages. It has been reported that the plastids of isogamous species are biparentally inherited whereas the plastids of oogamous species are maternally inherited. In contrast, in both isogamous and oogamous species, the mitochondria are usually inherited maternally. To investigate whether there is any relationship between the modes of inheritance and organellar genome architecture, we sequenced six plastid genomes (ptDNA) and two mitochondrial genomes (mtDNA) of isogamous species from the Ectocarpales and compared them with previously sequenced organellar genomes. We found that the biparentally inherited ptDNAs of isogamous species presented distinctive structural rearrangements whereas maternally inherited ptDNAs of oogamous species showed no rearrangements. Our analysis permits the hypothesis that structural rearrangements in ptDNAs may be a consequence of the mode of inheritance.

Differential gene expression in fronds and stolons of the siphonous macroalga, Caulerpa lentillifera.

The green alga, Caulerpa lentillifera, is composed of a single cell with multiple nuclei, but it possesses structures analogous to leaves or fronds, stems or stolons, and roots or rhizoids. To understand molecular mechanisms involved in formation and function of these structures, we carried out RNA-seq analysis of fronds and stolons (including rhizoids). Taking advantage of the decoded genome of C. lentillifera, the present RNA-seq analysis addressed transcripts corresponding to 9,311 genes identified in the genome. RNA-seq data suggested that 8,734 genes are expressed in sporophytes. Despite the siphonous body of the alga, differential gene expression was evident in the two structures. 1,027 (11.8%) and 1,129 (12.9%) genes were preferentially expressed in fronds and stolons, respectively, while the remaining 6,578 (75.3%) genes were expressed at the same level in both. Most genes preferentially expressed in fronds are associated with photosynthesis and plant hormone pathways, including abscisic acid signaling. In contrast, those preferentially expressed in stolons are associated with translation and DNA replication. These results indicate that gene expression is regulated differently between fronds and stolons, which probably governs the function of each structure. Together with genomic information, the present transcriptomic data provide genic information about development and physiology of this unique, siphonous organism.

A draft nuclear-genome assembly of the acoel flatworm Praesagittifera naikaiensis.

BACKGROUND: Acoels are primitive bilaterians with very simple soft bodies, in which many organs, including the gut, are not developed. They provide platforms for studying molecular and developmental mechanisms involved in the formation of the basic bilaterian body plan, whole-body regeneration, and symbiosis with photosynthetic microalgae. Because genomic information is essential for future research on acoel biology, we sequenced and assembled the nuclear genome of an acoel, Praesagittifera naikaiensis. FINDINGS: To avoid sequence contamination derived from symbiotic microalgae, DNA was extracted from embryos that were free of algae. More than 290x sequencing coverage was achieved using a combination of Illumina (paired-end and mate-pair libraries) and PacBio sequencing. RNA sequencing and Iso-Seq data from embryos, larvae, and adults were also obtained. First, a preliminary ∼17-kilobase pair (kb) mitochondrial genome was assembled, which was deleted from the nuclear sequence assembly. As a result, a draft nuclear genome assembly was ∼656 Mb in length, with a scaffold N50 of 117 kb and a contig N50 of 57 kb. Although ∼70% of the assembled sequences were likely composed of repetitive sequences that include DNA transposons and retrotransposons, the draft genome was estimated to contain 22,143 protein-coding genes, ∼99% of which were substantiated by corresponding transcripts. We could not find horizontally transferred microalgal genes in the acoel genome. Benchmarking Universal Single-Copy Orthologs analyses indicated that 77% of the conserved single-copy genes were complete. Pfam domain analyses provided a basic set of gene families for transcription factors and signaling molecules. CONCLUSIONS: Our present sequencing and assembly of the P. naikaiensis nuclear genome are comparable to those of other metazoan genomes, providing basic information for future studies of genic and genomic attributes of this animal group. Such studies may shed light on the origins and evolution of simple bilaterians.

A siphonous macroalgal genome suggests convergent functions of homeobox genes in algae and land plants.

Genome evolution and development of unicellular, multinucleate macroalgae (siphonous algae) are poorly known, although various multicellular organisms have been studied extensively. To understand macroalgal developmental evolution, we assembled the ∼26 Mb genome of a siphonous green alga, Caulerpa lentillifera, with high contiguity, containing 9,311 protein-coding genes. Molecular phylogeny using 107 nuclear genes indicates that the diversification of the class Ulvophyceae, including C. lentillifera, occurred before the split of the Chlorophyceae and Trebouxiophyceae. Compared with other green algae, the TALE superclass of homeobox genes, which expanded in land plants, shows a series of lineage-specific duplications in this siphonous macroalga. Plant hormone signalling components were also expanded in a lineage-specific manner. Expanded transport regulators, which show spatially different expression, suggest that the structural patterning strategy of a multinucleate cell depends on diversification of nuclear pore proteins. These results not only imply functional convergence of duplicated genes among green plants, but also provide insight into evolutionary roots of green plants. Based on the present results, we propose cellular and molecular mechanisms involved in the structural differentiation in the siphonous alga.