半索動物ヒメギボシムシ再生に関する進化発生学的研究

内容の要約広島大学(Hiroshima University)博士(理学)Doctor of Sciencedoctora

Xenacoelomorph-Specific Hox Peptides: Insights into the Phylogeny of Acoels, Nemertodermatids, and Xenoturbellids

Xenacoelomorpha has recently been proposed as an animal taxon that includes acoels, nemertodermatids, and xenoturbellids. Their flattened bodies are very simple and lack discrete organs. The Acoela and Nemertodermatida (which comprise Acoelomorpha) were traditionally regarded as early-diverged extant orders of the class Turbellaria of the phylum Platyhelminthes. Recent anatomical studies and molecular phylogenetic studies demonstrate that the two groups belong to the phylum Xenacoelomorpha together with Xenoturbellida. However, debate remains in regard to whether Xenacoelomorpha is monophyletic, and whether xenacoelomorphs are sisters to all other bilaterians or have close affinity to ambulacrarians. The present study addresses the first question by examining the presence or absence of diagnostic peptide sequences shared by the three taxa. Hox genes have been used to investigate the phylogenetic relationships of metazoans. It has been shown that lophotrochozoans, rotifers, and chaetognaths share diagnostic peptide sequences in the C-terminal region of the Lox5 (Hox5/6/7) homeodomain proteins, which supports the clustering of these taxa. Examination of the decoded genome of the acoel Praesagittifera naikaiensis and reported xenacoelomorph Hox genes revealed that acoels share a peptide NLK(S/T)MSQ(V/I)D, which starts immediately after the homeodomain sequence of the central Hox4/5/6. In addition, we found another diagnostic peptide, KEGKL, in the C-terminal region of the anterior Hox1, which is shared by all the three groups of xenacoelomorphs, but not other bilaterians. Furthermore, two acoels, Praesagittifera naikaiensis and Symsagittifera roscoffensis, share another peptide SG(A/P)-PGM in the posterior Hox9/11/13. These results support the designation of the phylum Xenacoelomorpha, in which Acoela is a discrete group

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

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

Draft genome of the brown alga, Nemacystus decipiens, Onna-1 strain: Fusion of genes involved in the sulfated fucan biosynthesis pathway

The brown alga, Nemacystus decipiens ("ito-mozuku" in Japanese), is one of the major edible seaweeds, cultivated principally in Okinawa, Japan. N. decipiens is also a significant source of fucoidan, which has various physiological activities. To facilitate brown algal studies, we decoded the ~154 Mbp draft genome of N. decipiens Onna-1 strain. The genome is estimated to contain 15,156 protein-coding genes, ~78% of which are substantiated by corresponding mRNAs. Mitochondrial genes analysis showed a close relationship between N. decipiens and Cladosiphon okamuranus. Comparisons with the C. okamuranus and Ectocarpus siliculosus genomes identified a set of N. decipiens-specific genes. Gene ontology annotation showed more than half of these are classified as molecular function, enzymatic activity, and/or biological process. Extracellular matrix analysis revealed domains shared among three brown algae. Characterization of genes that encode enzymes involved in the biosynthetic pathway for sulfated fucan showed two sets of genes fused in the genome. One is a fusion of L-fucokinase and GDP-fucose pyrophosphorylase genes, a feature shared with C. okamuranus. Another fusion is between an ST-domain-containing gene and an alpha/beta hydrolase gene. Although the function of fused genes should be examined in future, these results suggest that N. decipiens is another promising source of fucoidan

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

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

Integrated omics unveil the secondary metabolic landscape of a basal dinoflagellate

Abstract: 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

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

Chloroplast acquisition without the gene transfer in kleptoplastic sea slugs, Plakobranchus ocellatus

Some sea slugs sequester chloroplasts from algal food in their intestinal cells and photosynthesize for months. This phenomenon, kleptoplasty, poses a question of how the chloroplast retains its activity without the algal nucleus. There have been debates on the horizontal transfer of algal genes to the animal nucleus. To settle the arguments, this study reported the genome of a kleptoplastic sea slug, Plakobranchus ocellatus, and found no evidence of photosynthetic genes encoded on the nucleus. Nevertheless, it was confirmed that light illumination prolongs the life of mollusk under starvation. These data presented a paradigm that a complex adaptive trait, as typified by photosynthesis, can be transferred between eukaryotic kingdoms by a unique organelle transmission without nuclear gene transfer. Our phylogenomic analysis showed that genes for proteolysis and immunity undergo gene expansion and are up-regulated in chloroplast-enriched tissue, suggesting that these molluskan genes are involved in the phenotype acquisition without horizontal gene transfer