Ken-Ichiro Ishida

Publications of Ken-Ichiro Ishida

• Microheliella maris (Microhelida ord. n.), an Ultrastructurally Highly Distinctive New Axopodial Protist Species and Genus, and the Unity of Phylum Heliozoa.

  Authors: Akinori Yabuki, Ema E Chao, Ken-Ichiro Ishida, Thomas Cavalier-Smith

  Protist. 12/2011;

  A new heliozoan, Microheliella maris, has sufficiently distinctive ultrastructure to merit a new order, Microhelida. Its 18S and 28S rRNA genes were sequenced earlier under the informal name 'marineA new heliozoan, Microheliella maris, has sufficiently distinctive ultrastructure to merit a new order, Microhelida. Its 18S and 28S rRNA genes were sequenced earlier under the informal name 'marine microheliozoan'; we here sequenced its Hsp90 gene. A three-gene tree suggests that it is distantly related to centrohelids and others in chromist subkingdom Hacrobia; but it is too divergent to be placed accurately by few genes. Unlike centrohelids, its central spherical centrosome has two concentric granular shells and a dense core devoid of a trilaminar central disc. Microtubules radiate from the centrosomal shells. Unlike centrohelids, axopodia have only three microtubules, fixed basally by dense plasma membrane anchors, and bear terminal and lateral haptosome-like extrusomes. As in the heliomonad Heliomorpha, the centrosome is embedded in a nuclear cavity, and centrosomal microtubules traverse the nucleus inside cytoplasmic channels. A novel filogranular network interconnects mitochondria, ER, and plasma membrane. The microbody is attached to the nucleus and mitochondrion, which has vermicular tubular cristae. We group Microhelida and Heliomonadida, purged of dissimilar flagellates, as a new tubulicristate class Endohelea within phylum Heliozoa. Previously misassigned GenBank 18S rDNA sequences reveal Microhelida as diverse and ancient. We discuss principles underlying the biogenesis and diversity of axopodial patterns.

• Molecular genetic and chemotaxonomic characterization of the terrestrial cyanobacterium Nostoc commune and its neighboring species.

  Authors: Hiromi Arima, Noriomi Horiguchi, Shinichi Takaichi, Rumiko Kofuji, Ken-Ichiro Ishida, Keishiro Wada, Toshio Sakamoto

  FEMS microbiology ecology. 09/2011; 79(1):34-45.

  The phylogeny of the terrestrial cyanobacterium Nostoc commune and its neighboring Nostoc species was studied using molecular genetic and chemotaxonomic approaches. At least eight genotypes of N.The phylogeny of the terrestrial cyanobacterium Nostoc commune and its neighboring Nostoc species was studied using molecular genetic and chemotaxonomic approaches. At least eight genotypes of N. commune were characterized by the differences among 16S rRNA gene sequences and the petH gene encoding ferredoxin-NADP⁺ oxidoreductase and by random amplified polymorphic DNA analysis. The genotypes of N. commune were distributed in Japan without regional specificity. The nrtP gene encoding NrtP-type nitrate/nitrite permease was widely distributed in the genus Nostoc, suggesting that the occurrence of the nrtP gene can be one of the characteristic features that separate cyanobacteria into two groups. The wspA gene encoding a 36-kDa water stress protein was only found in N. commune and Nostoc verrucosum, suggesting that these Nostoc species that form massive colonies with extracellular polysaccharides can be exclusively characterized by the occurrence of the wspA gene. Fifteen species of Nostoc and Anabaena were investigated by comparing their carotenoid composition. Three groups with distinct patterns of carotenoids were related to the phylogenic tree constructed on the basis of 16S rRNA sequences. Nostoc commune and Nostoc punctiforme were clustered in one monophyletic group and characterized by the occurrence of nostoxanthin, canthaxanthin, and myxol glycosides.

• Tsukubamonas globosa n. gen., n. sp., a novel excavate flagellate possibly holding a key for the early evolution in "Discoba".

  Authors: Akinori Yabuki, Takeshi Nakayama, Naoji Yubuki, Tetsuo Hashimoto, Ken-Ichiro Ishida, Yuji Inagaki

  The Journal of eukaryotic microbiology. 05/2011; 58(4):319-31.

  We report the ultrastructure and phylogenetic position of a free-living heterotrophic flagellate, Tsukubamonas globosa n. gen., n. sp. This flagellate was isolated from a pond in the University ofWe report the ultrastructure and phylogenetic position of a free-living heterotrophic flagellate, Tsukubamonas globosa n. gen., n. sp. This flagellate was isolated from a pond in the University of Tsukuba, Japan. Under light microscopy, the spherical vegetative cells were naked and highly vacuolated, and always swam with rotating motion. Electron microscopic observations revealed that T. globosa possessed a ventral feeding groove, which is one of the hallmark characteristics of the supergroup Excavata. The position of T. globosa was unresolved in the small subunit ribosomal RNA phylogeny. On the other hand, a multigene phylogeny using α-tubulin, β-tubulin, actin, heat shock protein 90, and translation elongation factor 2 robustly united T. globosa with members of the "Discoba" clade of Excavata, composed of jakobids, euglenozoans, and heteroloboseans, although the precise position of T. globosa in this clade remained unresolved. Our detailed morphological comparisons elucidated that T. globosa possessed a novel set of morphological features, and could not be classified into any taxa in the Discoba clade. Instead we classified T. globosa into Tsukubamonadidae n. fam. under Tsukubamonadida n. ord.

• Mataza hastifera n. g., n. sp.: a possible new lineage in the Thecofilosea (Cercozoa).

  Authors: Akinori Yabuki, Ken-Ichiro Ishida

  The Journal of eukaryotic microbiology. 01/2011; 58(2):94-102.

  A new cercozoan flagellate Mataza hastifera n. g., n. sp. is described from a surface seawater sample collected in Tokyo Bay. Cells are 3-5 μm in diameter and have two flagella. The cells alternateA new cercozoan flagellate Mataza hastifera n. g., n. sp. is described from a surface seawater sample collected in Tokyo Bay. Cells are 3-5 μm in diameter and have two flagella. The cells alternate between swimming and stationary states in culture. Swimming cells have a nodding motion. Phylogenetic analyses using small subunit rDNA sequences demonstrate that M. hastifera belongs to the clade comprised of only environmental sequences closely related to thecofilosean cercozoans. Ultrastructural observations reveal that M. hastifera is quite similar to members of Cryomonadida, an order in Thecofilosea, and especially to Cryothecomonas spp. The cell of M. hastifera is covered with a thin double-layered theca and possesses a cylinder-shaped extrusome, as reported from cryomonads. On the other hand, the funnel that is characteristic of cryomonads was not found in the flagellar pit of M. hastifera. Combining both morphological and molecular analyses, we conclude that M. hastifera is a new lineage in Thecofilosea and suggest that Thecofilosea may be a larger group than previously thought.

• Spheroid bodies in rhopalodiacean diatoms were derived from a single endosymbiotic cyanobacterium.

  Authors: Takuro Nakayama, Yuko Ikegami, Takeshi Nakayama, Ken-Ichiro Ishida, Yuji Inagaki, Isao Inouye

  Journal of plant research. 01/2011; 124(1):93-7.

  Members of the diatom family rhopalodiaceae possess cyanobacteria-derived intracellular structures called spheroid bodies (SBs) that very likely carry out nitrogen fixation. Due to the shortage ofMembers of the diatom family rhopalodiaceae possess cyanobacteria-derived intracellular structures called spheroid bodies (SBs) that very likely carry out nitrogen fixation. Due to the shortage of molecular data from SBs and rhopalodiacean diatoms, it remains unclear how SBs were established and spread in rhopalodiacean diatoms. We here amplified the small subunit ribosomal DNA sequences from both host and SB in three rhopalodiacean diatom species, Epithemia turgida, E. sorex, and Rhopalodia gibba. Phylogenetic analyses considering these new sequences clearly indicate that the SBs were acquired by a common ancestor of rhopalodiacean diatoms and have been retained during host speciation.

• Internal plastid-targeting signal found in a RubisCO small subunit protein of a chlorarachniophyte alga.

  Authors: Yoshihisa Hirakawa, Ken-ichiro Ishida

  The Plant journal : for cell and molecular biology. 11/2010; 64(3):402-10.

  In all plants and algae, most plastid proteins are encoded by the nuclear genome and, consequently, need to be transported into plastids across multiple membranes. In organisms with secondaryIn all plants and algae, most plastid proteins are encoded by the nuclear genome and, consequently, need to be transported into plastids across multiple membranes. In organisms with secondary plastids, which evolved by secondary endosymbioses, and are surrounded by three or four envelope membranes, precursors of nuclear-encoded plastid proteins generally have an N-terminal bipartite targeting sequence that consists of an endoplasmic reticulum (ER)-targeting signal peptide (SP) and a transit peptide-like (TPL) sequence. The bipartite targeting sequences have been demonstrated to be necessary and sufficient for targeting proteins into the plastids of many algal groups, including chlorarachniophytes. Here, we report a new type of targeting signal that is required for delivering a RubisCO small subunit (RbcS) protein into the secondary plastids of chlorarachniophyte algae. In this study, we analyzed the plastid-targeting ability of an RbcS pre-protein, using green fluorescent protein (GFP) as a reporter molecule in chlorarachniophyte cells. We demonstrate that the N-terminal bipartite targeting sequence of the RbcS pre-protein is not sufficient, and that a part of the mature protein is also necessary for plastid targeting. By deletion analyses of amino acids, we determined the approximate location of an internal plastid-targeting signal within the mature protein, which is involved in targeting the protein from the ER into the chlorarachniophyte plastids.

• Cercozoa comprises both EF-1α-containing and EFL-containing members.

  Authors: Ryoma Kamikawa, Akinori Yabuki, Takuro Nakayama, Ken-ichiro Ishida, Tetsuo Hashimoto, Yuji Inagaki

  European journal of protistology. 11/2010; 47(1):24-8.

  Elongation factor 1α (EF-1α) and elongation factor-like protein (EFL) are considered to be functionally equivalent proteins involved in peptide synthesis. Eukaryotes can be fundamentally divided intoElongation factor 1α (EF-1α) and elongation factor-like protein (EFL) are considered to be functionally equivalent proteins involved in peptide synthesis. Eukaryotes can be fundamentally divided into 'EF-1α-containing' and 'EFL-containing' types. Recently, EF-1α and EFL genes have been surveyed across the diversity of eukaryotes to explore the origin and evolution of EFL genes. Although the phylum Cercozoa is a diverse group, gene data for either EFL or EF-1α are absent from all cercozoans except chlorarachniophytes which were previously defined as EFL-containing members. Our survey revealed that two members of the cercozoan subphylum Filosa (Thaumatomastix sp. and strain YPF610) are EFL-containing members. Importantly, we identified EF-1α genes from two members of Filosa (Paracercomonas marina and Paulinella chromatophora) and a member of the other subphylum Endomyxa (Filoreta japonica). All cercozoan EFL homologues could not be recovered as a monophyletic group in maximum-likelihood and Bayesian analyses, suggesting that lateral gene transfer was involved in the EFL evolution in this protist assemblage. In contrast, EF-1α analysis successfully recovered a monophyly of three homologues sampled from the two cercozoan subphyla. Based on the results, we postulate that cercozoan EF-1α genes have been vertically inherited, and the current EFL-containing species may have secondarily lost their EF-1α genes.

• Characterization of periplastidal compartment-targeting signals in chlorarachniophytes.

  Authors: Yoshihisa Hirakawa, Gillian H Gile, Shuhei Ota, Patrick J Keeling, Ken-ichiro Ishida

  Molecular biology and evolution. 07/2010; 27(7):1538-45.

  Secondary plastids are acquired by the engulfment and retention of eukaryotic algae, which results in an additional surrounding membrane or pair of membranes relative to the more familiar primarySecondary plastids are acquired by the engulfment and retention of eukaryotic algae, which results in an additional surrounding membrane or pair of membranes relative to the more familiar primary plastids of land plants. In most cases, the endocytosed alga loses its eukaryotic genome as it becomes integrated, but in two algal groups, the cryptophytes and chlorarachniophytes, the secondary plastids retain a vestigial nucleus in the periplastidal compartment (PPC), the remnant eukaryotic cytoplasm between the inner and the outer membrane pairs. Many essential housekeeping genes are missing from these reduced genomes, suggesting that they are now encoded in the host nucleus and their products are targeted to the PPC. One such nucleus-encoded, PPC-targeted protein, the translation elongation factor like (EFL) was recently identified in chlorarachniophytes. It bears an N-terminal-targeting sequence comprising a signal peptide and a transit peptide-like sequence (TPL) similar to the plastid-targeted proteins of chlorarachniophytes as well as a hydrophilic C-terminal extension rich in lysine and aspartic acid. Here, we characterize the function of the N- and C-terminal extensions of PPC-targeted EFL in transformed chlorarachniophyte cells. Using green fluorescent protein as a reporter molecule, we demonstrate that several negatively charged amino acids within the TPL are essential for accurate targeting to the PPC. Our findings further reveal that the C-terminal extension functions as a PPC retention signal in combination with an N-terminal plastid-targeting peptide, which suggests that plastid and PPC proteins may be sorted in the PPC.

• Differential gene retention in plastids of common recent origin.

  Authors: Adrian Reyes-Prieto, Hwan Su Yoon, Ahmed Moustafa, Eun Chan Yang, Robert A Andersen, Sung Min Boo, Takuro Nakayama, Ken-ichiro Ishida, Debashish Bhattacharya

  Molecular biology and evolution. 07/2010; 27(7):1530-7.

  The cyanobacterium-derived plastids of algae and plants have supported the diversification of much of extant eukaryotic life. Inferences about early events in plastid evolution must rely onThe cyanobacterium-derived plastids of algae and plants have supported the diversification of much of extant eukaryotic life. Inferences about early events in plastid evolution must rely on reconstructing events that occurred over a billion years ago. In contrast, the photosynthetic amoeba Paulinella chromatophora provides an exceptional model to study organelle evolution in a prokaryote-eukaryote (primary) endosymbiosis that occurred approximately 60 mya. Here we sequenced the plastid genome (0.977 Mb) from the recently described Paulinella FK01 and compared the sequence with the existing data from the sister taxon Paulinella M0880/a. Alignment of the two plastid genomes shows significant conservation of gene order and only a handful of minor gene rearrangements. Analysis of gene content reveals 66 differential gene losses that appear to be outright gene deletions rather than endosymbiotic gene transfers to the host nuclear genome. Phylogenomic analysis validates the plastid ancestor as a member of the Synechococcus-Prochlorococcus group, and the cyanobacterial provenance of all plastid genes suggests that these organelles were not targets of interphylum gene transfers after endosymbiosis. Inspection of 681 DNA alignments of protein-encoding genes shows that the vast majority have dN/dS ratios <1, providing evidence for purifying selection. Our study demonstrates that plastid genomes in sister taxa are strongly constrained by selection but follow distinct trajectories during the earlier phases of organelle evolution.

• Palpitomonas bilix gen. et sp. nov.: A novel deep-branching heterotroph possibly related to Archaeplastida or Hacrobia.

  Authors: Akinori Yabuki, Yuji Inagaki, Ken-ichiro Ishida

  Protist. 04/2010; 161(4):523-38.

  We describe the ultrastructure and putative molecular phylogenetic position of a free-living heterotrophic flagellate Palpitomonas bilix gen. et sp. nov. This flagellate is 3-8mum in size, andWe describe the ultrastructure and putative molecular phylogenetic position of a free-living heterotrophic flagellate Palpitomonas bilix gen. et sp. nov. This flagellate is 3-8mum in size, and possesses two subequal flagella, approximately 20mum long. Electron microscopical observations revealed that the flagellar apparatus of P. bilix resembles that of members of the green algal class, Charophyceae, while the mastigonemes of the P. bilix flagellum share some characteristics with those found in cryptophytes and telonemids. In order to better understand the phylogenetic position of P. bilix, we sequenced six commonly used phylogenetic marker genes encoding the small and large subunits of ribosomal RNA, alpha-tubulin, beta-tubulin, 90kDa heat shock protein, and translation elongation factor 2. Depending on the genes analyzed, P. bilix shows a generally weak phylogenetic affinity to either the newly erected Hacrobia, which includes cryptophytes and haptophytes, or to Archaeplastida. Since the current study identified no clear close relative of P. bilix, this novel biflagellate was classified into a new genus Palpitomonas with the higher-level classification being unclear. On the basis of the results of both morphological and molecular studies, we discuss the possibility that P. bilix might provide key information on the early evolution of major groups of photosynthetic eukaryotes.

• Protein targeting into secondary plastids of chlorarachniophytes.

  Authors: Yoshihisa Hirakawa, Kisaburo Nagamune, Ken-Ichiro Ishida

  Proceedings of the National Academy of Sciences of the United States of America. 08/2009;

  Most plastid proteins are encoded by the nuclear genome, and consequently, need to be transported into plastids across multiple envelope membranes. In diverse organisms possessing secondary plastids,Most plastid proteins are encoded by the nuclear genome, and consequently, need to be transported into plastids across multiple envelope membranes. In diverse organisms possessing secondary plastids, nuclear-encoded plastid precursor proteins (preproteins) commonly have an N-terminal extension that consists of an endoplasmic reticulum (ER)-targeting signal peptide and a transit peptide-like sequence (TPL). This bipartite targeting peptide is believed to be necessary for targeting the preproteins into the secondary plastids. Here, we newly demonstrate the function of the bipartite targeting peptides of an algal group, chlorarachniophytes, and characterize the functional domains of the TPL in the precursor of a plastid protein, ATP synthase delta subunit (AtpD), using a GFP as a reporter molecule. We show that the C-terminal portion of the TPL is important for targeting the AtpD preprotein from the ER into the chlorarachniophyte plastids, and several positively charged amino acids in the TPL are also necessary for transporting the preprotein across the 2 innermost plastid membranes. Compared with other groups with secondary plastids, the TPL functional domains of the chlorarachniophytes are unique, which might be caused by independent acquisition of their plastids.

• Another acquisition of a primary photosynthetic organelle is underway in Paulinella chromatophora.

  Authors: Takuro Nakayama, Ken-Ichiro Ishida

  Current biology : CB. 05/2009; 19(7):R284-5.

• A single origin of the photosynthetic organelle in different Paulinella lineages.

  Authors: Hwan Su Yoon, Takuro Nakayama, Adrian Reyes-Prieto, Robert A Andersen, Sung Min Boo, Ken-Ichiro Ishida, Debashish Bhattacharya

  BMC evolutionary biology. 02/2009; 9:98.

  BACKGROUND: Gaining the ability to photosynthesize was a key event in eukaryotic evolution because algae and plants form the base of the food chain on our planet. The eukaryotic machines ofBACKGROUND: Gaining the ability to photosynthesize was a key event in eukaryotic evolution because algae and plants form the base of the food chain on our planet. The eukaryotic machines of photosynthesis are plastids (e.g., chloroplast in plants) that evolved from cyanobacteria through primary endosymbiosis. Our knowledge of plastid evolution, however, remains limited because the primary endosymbiosis occurred more than a billion years ago. In this context, the thecate "green amoeba" Paulinella chromatophora is remarkable because it very recently (i.e., minimum age of approximately 60 million years ago) acquired a photosynthetic organelle (termed a "chromatophore"; i.e., plastid) via an independent primary endosymbiosis involving a Prochlorococcus or Synechococcus-like cyanobacterium. All data regarding P. chromatophora stem from a single isolate from Germany (strain M0880/a). Here we brought into culture a novel photosynthetic Paulinella strain (FK01) and generated molecular sequence data from these cells and from four different cell samples, all isolated from freshwater habitats in Japan. Our study had two aims. The first was to compare and contrast cell ultrastructure of the M0880/a and FK01 strains using scanning electron microscopy. The second was to assess the phylogenetic diversity of photosynthetic Paulinella to test the hypothesis they share a vertically inherited plastid that originated in their common ancestor. RESULTS: Comparative morphological analyses show that Paulinella FK01 cells are smaller than M0880/a and differ with respect to the number of scales per column. There are more distinctive, multiple fine pores on the external surface of FK01 than in M0880/a. Molecular phylogenetic analyses using multiple gene markers demonstrate these strains are genetically distinct and likely comprise separate species. The well-supported monophyly of the Paulinella chromatophora strains analyzed here using plastid-encoded 16S rRNA suggests strongly that they all share a common photosynthetic ancestor. The strain M0880/a is most closely related to Japanese isolates (Kanazawa-1, -2, and Kaga), whereas FK01 groups closely with a Kawaguchi isolate. CONCLUSION: Our results indicate that Paulinella chromatophora comprises at least two distinct evolutionary lineages and likely encompasses a broader taxonomic diversity than previously thought. The finding of a single plastid origin for both lineages shows these taxa to be valuable models for studying post-endosymbiotic cell and genome evolution.

• A single origin of the photosynthetic organelle in different Paulinella lineages

  Authors: Yoon Hwan Su, Takuro Nakayama, Adrian Reyes-Prieto, Robert Andersen, Boo Sung Min, Ken-Ichiro Ishida, Debashish Bhattacharya

  BMC Evolutionary Biology. 01/2009;

  Abstract Background Gaining the ability to photosynthesize was a key event in eukaryotic evolution because algae and plants form the base of the food chain on our planet. The eukaryotic machines ofAbstract Background Gaining the ability to photosynthesize was a key event in eukaryotic evolution because algae and plants form the base of the food chain on our planet. The eukaryotic machines of photosynthesis are plastids (e.g., chloroplast in plants) that evolved from cyanobacteria through primary endosymbiosis. Our knowledge of plastid evolution, however, remains limited because the primary endosymbiosis occurred more than a billion years ago. In this context, the thecate "green amoeba" Paulinella chromatophora is remarkable because it very recently (i.e., minimum age of ≈ 60 million years ago) acquired a photosynthetic organelle (termed a "chromatophore"; i.e., plastid) via an independent primary endosymbiosis involving a Prochlorococcus or Synechococcus -like cyanobacterium. All data regarding P. chromatophora stem from a single isolate from Germany (strain M0880/a). Here we brought into culture a novel photosynthetic Paulinella strain (FK01) and generated molecular sequence data from these cells and from four different cell samples, all isolated from freshwater habitats in Japan. Our study had two aims. The first was to compare and contrast cell ultrastructure of the M0880/a and FK01 strains using scanning electron microscopy. The second was to assess the phylogenetic diversity of photosynthetic Paulinella to test the hypothesis they share a vertically inherited plastid that originated in their common ancestor. Results Comparative morphological analyses show that Paulinella FK01 cells are smaller than M0880/a and differ with respect to the number of scales per column. There are more distinctive, multiple fine pores on the external surface of FK01 than in M0880/a. Molecular phylogenetic analyses using multiple gene markers demonstrate these strains are genetically distinct and likely comprise separate species. The well-supported monophyly of the Paulinella chromatophora strains analyzed here using plastid-encoded 16S rRNA suggests strongly that they all share a common photosynthetic ancestor. The strain M0880/a is most closely related to Japanese isolates (Kanazawa-1, -2, and Kaga), whereas FK01 groups closely with a Kawaguchi isolate. Conclusion Our results indicate that Paulinella chromatophora comprises at least two distinct evolutionary lineages and likely encompasses a broader taxonomic diversity than previously thought. The finding of a single plastid origin for both lineages shows these taxa to be valuable models for studying post-endosymbiotic cell and genome evolution.

• Partenskyella glossopodia gen. et sp. nov., the First Report of a Chlorarachniophyte that Lacks a Pyrenoid.

  Authors: Shuhei Ota, Daniel Vaulot, Florence Le Gall, Akinori Yabuki, Ken-Ichiro Ishida

  Protist. 12/2008;

  A new chlorarachniophyte, Partenskyella glossopodia gen. et sp. nov., is described from a culture isolated from the Mediterranean Sea pelagic waters and maintained as strain RCC365 at the RoscoffA new chlorarachniophyte, Partenskyella glossopodia gen. et sp. nov., is described from a culture isolated from the Mediterranean Sea pelagic waters and maintained as strain RCC365 at the Roscoff Culture Collection (France). Vegetative cells of P. glossopodia are non-motile naked spherical cells. However, flagellate and amoeboid stages are also present in its life cycle. The cells are 2-4mum in diameter containing a pale-green, cup-shaped chloroplast, 1-2 mitochondria, a nucleus, and a Golgi apparatus. Vesicles containing storage product-like material are also present. The chloroplast is surrounded by four membranes possessing a nucleomorph in the periplastidal compartment. The minute cell size and the absence of a pyrenoid at any stage of the life cycle are unique characteristics among the chlorarachniophytes, which justifies our proposition for a new genus for strain RCC365.

• Phylogeny of Novel Naked Filose and Reticulose Cercozoa: Granofilosea cl. n. and Proteomyxidea Revised.

  Authors: David Bass, Ema E-Y Chao, Sergey Nikolaev, Akinori Yabuki, Ken-Ichiro Ishida, Cédric Berney, Ursula Pakzad, Claudia Wylezich, Thomas Cavalier-Smith

  Protist. 11/2008;

  Naked filose and reticulose protozoa were long lumped as proteomyxids or left outside higher groups. We cultivated eight naked filose or reticulose strains, did light microscopy, 18S rDNA sequencingNaked filose and reticulose protozoa were long lumped as proteomyxids or left outside higher groups. We cultivated eight naked filose or reticulose strains, did light microscopy, 18S rDNA sequencing and phylogeny (showing all are Cercozoa), and sequenced 80 environmental 18S-types. Filose species belong in subphylum Filosa and reticulose ones in subphylum Endomyxa, making proteomyxids polyphyletic. We therefore transfer the classically mainly reticulose Proteomyxidea to Endomyxa, removing evident filosans as new class Granofilosea (including Desmothoracida, Acinetactis and new heliomonad family Heliomorphidae (new genus Heliomorpha (=Dimorpha)). Five new species of Limnofila gen. n. (L. mylnikovi; L. anglica; L. longa; L. oxoniensis; L. borokensis, previously misidentified as Biomyxa (=Gymnophrys) cometa) form a large freshwater clade (new order Limnofilida). Mesofila limnetica gen., sp. n. and Nanofila marina gen., sp. n. group separately in Granofilosea (Cryptofilida ord. n.). In Endomyxa, a new genus of reticulose proteomyxids (Filoreta marina, F. japonica, F. turcica spp. n., F. (=Corallomyxa) tenera comb. n.) forms a clade (Reticulosida) related to Gromiidea/Ascetosporea. Platyreta germanica gen., sp. n. and Arachnula impatiens are related vampyrellids (Aconchulinida) within a large clade beside Phytomyxea. Biomyxidae and Rhizoplasmidae fam. n. remain incertae sedis within Proteomyxidea. Gymnophrydium and Borkovia are revised. The reticulose Corallomyxa are unlike Filoreta and possibly Amoebozoa, not Cercozoa.

• Origins of plastids and glyceraldehyde-3-phosphate dehydrogenase genes in the green-colored dinoflagellate Lepidodinium chlorophorum.

  Authors: Kiyotaka Takishita, Masanobu Kawachi, Mary-Hélène Noël, Takuya Matsumoto, Natsuki Kakizoe, Makoto M Watanabe, Isao Inouye, Ken-Ichiro Ishida, Tetsuo Hashimoto, Yuji Inagaki

  Gene. 03/2008; 410(1):26-36.

  The dinoflagellate Lepidodinium chlorophorum possesses "green" plastids containing chlorophylls a and b (Chl a+b), unlike most dinoflagellate plastids with Chl a+c plus a carotenoid peridininThe dinoflagellate Lepidodinium chlorophorum possesses "green" plastids containing chlorophylls a and b (Chl a+b), unlike most dinoflagellate plastids with Chl a+c plus a carotenoid peridinin (peridinin-containing plastids). In the present study we determined 8 plastid-encoded genes from Lepidodinium to investigate the origin of the Chl a+b-containing dinoflagellate plastids. The plastid-encoded gene phylogeny clearly showed that Lepidodinium plastids were derived from a member of Chlorophyta, consistent with pigment composition. We also isolated three different glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes from Lepidodinium-one encoding the putative cytosolic "GapC" enzyme and the remaining two showing affinities to the "plastid-targeted GapC" genes. In a GAPDH phylogeny, one of the plastid-targeted GapC-like sequences robustly grouped with those of dinoflagellates bearing peridinin-containing plastids, while the other was nested in a clade of the homologues of haptophytes and dinoflagellate genera Karenia and Karlodinium bearing "haptophyte-derived" plastids. Since neither host nor plastid phylogeny suggested an evolutionary connection between Lepidodinium and Karenia/Karlodinium, a lateral transfer of a plastid-targeted GapC gene most likely took place from a haptophyte or a dinoflagellate with haptophyte-derived plastids to Lepidodinium. The plastid-targeted GapC data can be considered as an evidence for the single origin of plastids in haptophytes, cryptophytes, stramenopiles, and alveolates. However, in the light of Lepidodinium GAPDH data, we need to closely examine whether the monophyly of the plastids in the above lineages inferred from plastid-targeted GapC genes truly reflects that of the host lineages.

• Norrisiella sphaerica gen. et sp. nov., a new coccoid chlorarachniophyte from Baja California, Mexico.

  Authors: Shuhei Ota, Kunihiko Ueda, Ken-Ichiro Ishida

  Journal of plant research. 12/2007; 120(6):661-70.

  A new chlorarachniophyte, Norrisiella sphaerica S. Ota et K. Ishida gen. et sp. nov., from the coast of Baja California, Mexico is described. We examined its morphology, ultrastructure, and lifeA new chlorarachniophyte, Norrisiella sphaerica S. Ota et K. Ishida gen. et sp. nov., from the coast of Baja California, Mexico is described. We examined its morphology, ultrastructure, and life cycle in detail, using light microscopy, transmission electron microscopy, and time-lapse videomicroscopy. We found that this chlorarachniophyte possessed the following characteristics: (1) vegetative cells were coccoid and possessed a cell wall, (2) a pyrenoid was slightly invaded by plate-like periplastidial compartment from the tip of the pyrenoid, (3) a nucleomorph was located near the pyrenoid base in the periplastidial compartment, (4) cells reproduced vegetatively via autospores, and (5) a flagellate stage was present in the life cycle. This combination of characteristics differs from any of the described chlorarachniophyte genera, and therefore a new genus is established. Fluorescent microscopic observations suggested that the alga formed multinucleate cells prior to forming autospores. Time-lapse observations during autospore formation showed that cytokinesis occurred simultaneously in the multinucleate cells. Zoospores were also produced, and video sequences captured the release of zoospores from coccoid cells.

• Identification and transcription of transfer RNA genes in dinoflagellate plastid minicircles.

  Authors: Martha J Nelson, Yunkun Dang, Elena Filek, Zhaoduo Zhang, Vionnie Wing Chi Yu, Ken-Ichiro Ishida, Beverley R Green

  Gene. 06/2007; 392(1-2):291-8.

  The dinoflagellate chloroplast genome is unique in that the genes are found on small circular DNA molecules carrying from one to three genes. In addition, only 14 of the typical chloroplast-locatedThe dinoflagellate chloroplast genome is unique in that the genes are found on small circular DNA molecules carrying from one to three genes. In addition, only 14 of the typical chloroplast-located genes have so far been discovered on minicircles, while a number have been transferred to the nucleus. We have sequenced four new minicircles from the dinoflagellate Heterocapsa triquetra, three of which carry a single protein-coding gene (psbD, psbE, petD) and one that appears to be an "empty" circle. Using the tRNA prediction programs ARAGORN and tRNAscan-SE, tRNA-Met was found in the petD circle immediately downstream of the end of petD, while tRNA-Trp and tRNA-Pro were detected in the psbE and petD circles as well as in several chimeric circles of H. triquetra and the psbA minicircles of Heterocapsa pygmaea. RT-PCR showed that the tRNAs were co-transcribed with the protein-coding genes that preceded them, and cleaved from the precursor before a poly(U) tail was added to the mRNA.

• Protein targeting into plastids: a key to understanding the symbiogenetic acquisitions of plastids.

  Authors: Ken-ichiro Ishida

  Journal of plant research. 09/2005; 118(4):237-45.

  Recent progress in molecular phylogenetics has proven that photosynthetic eukaryotes acquired plastids via primary and secondary endosymbiosis and has given us information about the origin of eachRecent progress in molecular phylogenetics has proven that photosynthetic eukaryotes acquired plastids via primary and secondary endosymbiosis and has given us information about the origin of each plastid. How a photosynthetic endosymbiont became a plastid in each group is, however, poorly understood, especially for the organisms with secondary plastids. Investigating how a nuclear-encoded plastid protein is targeted into a plastid in each photosynthetic group is one of the most important keys to understanding the evolutionary process of symbiogenetic plastid acquisition and its diversity. For organisms which originated through primary endosymbiosis, protein targeting into plastids has been well studied at the molecular level. For organisms which originated through secondary endosymbiosis, molecular-level studies have just started on the plastid-targeted protein-precursor sequences and the targeting pathways of the precursors. However, little information is available about how the proteins get across the inner two or three envelope membranes in organisms with secondary plastids. A good in vitro protein-import system for isolated plastids and a cell transformation system must be established for each group of photosynthetic eukaryotes in order to understand the mechanisms, the evolutionary processes and the diversity of symbiogenetic plastid acquisitions in photosynthetic eukaryotes.