The Experts below are selected from a list of 315 Experts worldwide ranked by ideXlab platform
Huan Zhang - One of the best experts on this subject based on the ideXlab platform.
-
Nuclear Gene Transformation in the Dinoflagellate Oxyrrhis marina.
Microorganisms, 2020Co-Authors: Brittany N. Sprecher, Huan ZhangAbstract:The lack of a robust gene transformation tool that allows proper expression of foreign genes and functional testing for the vast number of nuclear genes in Dinoflagellates has greatly hampered our understanding of the fundamental biology in this ecologically important and evolutionarily unique lineage of microeukaryotes. Here, we report the development of a Dinoflagellate expression vector containing various DNA elements from phylogenetically separate Dinoflagellate lineages, an electroporation protocol, and successful expression of introduced genes in an early branching Dinoflagellate, Oxyrrhis marina. This protocol, involving the use of Lonza’s Nucleofector and a codon-optimized antibiotic resistance gene, has been successfully used to produce consistent results in several independent experiments for O. marina. It is anticipated that this protocol will be adaptable for other Dinoflagellates and will allow characterization of many novel Dinoflagellate genes.
-
Nuclear gene transformation in a Dinoflagellate
bioRxiv, 2019Co-Authors: Brittany N. Sprecher, Huan ZhangAbstract:ABSTRACT The lack of a robust gene transformation tool that allows functional testing of the vast number of nuclear genes in Dinoflagellates has greatly hampered our understanding of fundamental biology in this ecologically important and evolutionarily unique lineage. Here we report the development of a Dinoflagellate expression vector, an electroporation protocol, and successful expression of introduced genes in the Dinoflagellate Oxyrrhis marina. This protocol, involving the use of Lonza’s Nucleofector and a codon optimized antibiotic resistance gene, has been successfully used to produce consistent results in several independent experiments. It is anticipated that this protocol will be adaptable for other Dinoflagellates and will allow characterization of many novel Dinoflagellate genes.
-
high level diversity of Dinoflagellates in the natural environment revealed by assessment of mitochondrial cox1 and cob genes for Dinoflagellate dna barcoding
Applied and Environmental Microbiology, 2009Co-Authors: Huan Zhang, Yunyun Zhuang, Lilibeth MirandaAbstract:DNA barcoding is a diagnostic technique for species identification using a short, standardized DNA. An effective DNA barcoding marker would be very helpful for unraveling the poorly understood species diversity of Dinoflagellates in the natural environment. In this study, the potential utility for DNA barcoding of mitochondrial cytochrome c oxidase 1 (cox1) and cytochrome b (cob) was assessed. Among several primer sets examined, the one amplifying a 385-bp cob fragment was most effective for Dinoflagellates. This short cob fragment is easy to sequence and yet possess reasonable taxon resolution. While the lack of a uniform gap between interspecific and intraspecific distances poses difficulties in establishing a phylum-wide species-discriminating distance threshold, the variability of cob allows recognition of species within particular lineages. The potential of this cob fragment as a Dinoflagellate species marker was further tested by applying it to an analysis of the Dinoflagellate assemblages in Long Island Sound (LIS) and Mirror Lake in Connecticut. In LIS, a highly diverse assemblage of Dinoflagellates was detected. Some taxa can be identified to the species and some to the genus level, including a taxon distinctly related to the bipolar species Polarella glacialis, and the large number of others cannot be clearly identified, due to the inadequate database. In Mirror Lake, a Ceratium species and an unresolved taxon were detected, exhibiting a temporal transition from one to the other. We demonstrate that this 385-bp cob fragment is promising for lineage-wise Dinoflagellate species identification, given an adequate database.
-
Retrieval of missing spliced leader in Dinoflagellates.
PLOS ONE, 2009Co-Authors: Huan ZhangAbstract:: Spliced leader (SL) trans-splicing has recently been shown to be a common mRNA processing mechanism in Dinoflagellates, in which a short (22-nt) sequence, DCCGUAGCCAUUUUGGCUCAAG (D = U, A, or G), is transplanted from the 5'-end of a small non-coding RNA (SL RNA) to the 5' end of mRNA molecules. The widespread existence of the mechanism in Dinoflagellates has been demonstrated by detection of this SL (DinoSL) in a wide phylogenetic range of Dinoflagellates. Furthermore, the presence of DinoSL in the transcripts of highly diverse groups of nuclear-encoded genes has led us to postulate that SL trans-splicing is universal in Dinoflagellate nuclear genome. However, some observations inconsistent to this postulation have been reported, exemplified by a recent article reporting apparent absence of DinoSL in the transcripts of some nuclear-encoded genes in Amphidinium carterae. Absence of SL in these gene transcripts would have important implication on gene regulation in Dinoflagellates and utility of DinoSL as a universal Dinoflagellate-specific primer to study Dinoflagellate transcriptomics. In this study, we re-examined transcripts of these genes and found that all of them actually contained DinoSL. Therefore, results to date are consistent to our initial postulation that DinoSL occurs in all Dinoflagellate nuclear-encoded mRNAs.
-
phylogeny of Dinoflagellates based on mitochondrial cytochrome b and nuclear small subunit rdna sequence comparisons
Journal of Phycology, 2005Co-Authors: Huan Zhang, Debashish BhattacharyaAbstract:Despite their evolutionary and ecological importance, Dinoflagellate phylogeny remains poorly resolved. Here we explored the utility of mitochondrial cytochrome b (cob) in inferring a Dinoflagellate tree and focused on resolving the relationship between fucoxanthin-and peridinin-containing taxa. Trees were inferred using cob and small subunit rDNA alone or in combination as concatenated data and including members of the six major Dinoflagellate orders. Many regions of the cob DNA or protein and rDNA trees were congruent with support for the monophyly of Symbiodinium spp. Freudenthal and of the Prorocentrales and the early divergence of Crypthecodinium cohnii Seligo in Grasse. However, these markers provided differing support for the monophyly of Pfiesteria spp. Steidinger et Burkholder (only supported strongly by rDNA) and of the fucoxanthin Dinoflagellates with Akashiwo sp. (Hirasaka) Hansen et Moestrup (Gymnodiniales, only supported strongly by the cob data). The approximately unbiased (AU) test was used to assess these results using 13-and 11-taxon (excluding apicomplexans) backbone maximum likelihood trees inferred from the combined cob + rDNA data. The AU test suggested that our data were insufficient to resolve the phylogenetic position of Symbiodinium spp. and that the ancestral position of C. cohnii might have resulted from long-branch attraction to the apicomplexan outgroup. We found significant support, however, for the association of fucoxanthin Dinoflagellates with Akashiwo sp. The monophyly and relatively derived position of the Gymnodiniales in our cob DNA and protein trees and in the cob +rDNA tree is consistent with the tertiary endosymbiotic origin of the plastid in fucoxanthin Dinoflagellates.
Thomas E Decoursey - One of the best experts on this subject based on the ideXlab platform.
-
voltage gated proton channel in a Dinoflagellate
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Susan M.e. Smith, Deri Morgan, Boris Musset, Vladimir V Cherny, Allen R Place, Woodland J Hastings, Thomas E DecourseyAbstract:Fogel and Hastings first hypothesized the existence of voltage-gated proton channels in 1972 in bioluminescent Dinoflagellates, where they were thought to trigger the flash by activating luciferase. Proton channel genes were subsequently identified in human, mouse, and Ciona intestinalis, but their existence in Dinoflagellates remained unconfirmed. We identified a candidate proton channel gene from a Karlodinium veneficum cDNA library based on homology with known proton channel genes. K. veneficum is a predatory, nonbioluminescent Dinoflagellate that produces toxins responsible for fish kills worldwide. Patch clamp studies on the heterologously expressed gene confirm that it codes for a genuine voltage-gated proton channel, kHV1: it is proton-specific and activated by depolarization, its gH–V relationship shifts with changes in external or internal pH, and mutation of the selectivity filter (which we identify as Asp51) results in loss of proton-specific conduction. Indirect evidence suggests that kHV1 is monomeric, unlike other proton channels. Furthermore, kHV1 differs from all known proton channels in activating well negative to the Nernst potential for protons, EH. This unique voltage dependence makes the Dinoflagellate proton channel ideally suited to mediate the proton influx postulated to trigger bioluminescence. In contrast to vertebrate proton channels, whose main function is acid extrusion, we propose that proton channels in Dinoflagellates have fundamentally different functions of signaling and excitability.
Susan M.e. Smith - One of the best experts on this subject based on the ideXlab platform.
-
Hv1 Proton Channels in Dinoflagellates: Not Just for Bioluminescence?
Journal of Eukaryotic Microbiology, 2018Co-Authors: Gabriel Kigundu, Jennifer L. Cooper, Susan M.e. SmithAbstract:: Bioluminescence in Dinoflagellates is controlled by HV 1 proton channels. Database searches of Dinoflagellate transcriptomes and genomes yielded hits with sequence features diagnostic of all confirmed HV 1, and show that HV 1 is widely distributed in the Dinoflagellate phylogeny including the basal species Oxyrrhis marina. Multiple sequence alignments followed by phylogenetic analysis revealed three major subfamilies of HV 1 that do not correlate with presence of theca, autotrophy, geographic location, or bioluminescence. These data suggest that most Dinoflagellates express a HV 1 which has a function separate from bioluminescence. Sequence evidence also suggests that Dinoflagellates can contain more than one HV 1 gene.
-
voltage gated proton channel in a Dinoflagellate
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Susan M.e. Smith, Deri Morgan, Boris Musset, Vladimir V Cherny, Allen R Place, Woodland J Hastings, Thomas E DecourseyAbstract:Fogel and Hastings first hypothesized the existence of voltage-gated proton channels in 1972 in bioluminescent Dinoflagellates, where they were thought to trigger the flash by activating luciferase. Proton channel genes were subsequently identified in human, mouse, and Ciona intestinalis, but their existence in Dinoflagellates remained unconfirmed. We identified a candidate proton channel gene from a Karlodinium veneficum cDNA library based on homology with known proton channel genes. K. veneficum is a predatory, nonbioluminescent Dinoflagellate that produces toxins responsible for fish kills worldwide. Patch clamp studies on the heterologously expressed gene confirm that it codes for a genuine voltage-gated proton channel, kHV1: it is proton-specific and activated by depolarization, its gH–V relationship shifts with changes in external or internal pH, and mutation of the selectivity filter (which we identify as Asp51) results in loss of proton-specific conduction. Indirect evidence suggests that kHV1 is monomeric, unlike other proton channels. Furthermore, kHV1 differs from all known proton channels in activating well negative to the Nernst potential for protons, EH. This unique voltage dependence makes the Dinoflagellate proton channel ideally suited to mediate the proton influx postulated to trigger bioluminescence. In contrast to vertebrate proton channels, whose main function is acid extrusion, we propose that proton channels in Dinoflagellates have fundamentally different functions of signaling and excitability.
Patrick J. Keeling - One of the best experts on this subject based on the ideXlab platform.
-
microbial arms race ballistic nematocysts in Dinoflagellates represent a new extreme in organelle complexity
Science Advances, 2017Co-Authors: Gregory S Gavelis, Kevin C Wakeman, Urban Tillmann, Christina Ripken, Satoshi Mitarai, Maria Herranz, Suat Ozbek, Thomas W Holstein, Patrick J. KeelingAbstract:We examine the origin of harpoon-like secretory organelles (nematocysts) in Dinoflagellate protists. These ballistic organelles have been hypothesized to be homologous to similarly complex structures in animals (cnidarians); but we show, using structural, functional, and phylogenomic data, that nematocysts evolved independently in both lineages. We also recorded the first high-resolution videos of nematocyst discharge in Dinoflagellates. Unexpectedly, our data suggest that different types of Dinoflagellate nematocysts use two fundamentally different types of ballistic mechanisms: one type relies on a single pressurized capsule for propulsion, whereas the other type launches 11 to 15 projectiles from an arrangement similar to a Gatling gun. Despite their radical structural differences, these nematocysts share a single origin within Dinoflagellates and both potentially use a contraction-based mechanism to generate ballistic force. The diversity of traits in Dinoflagellate nematocysts demonstrates a stepwise route by which simple secretory structures diversified to yield elaborate subcellular weaponry.
-
Molecular data and the evolutionary history of Dinoflagellates
European Journal of Protistology, 2004Co-Authors: Juan F. Saldarriaga, F. J. R. “max” Taylor, Thomas Cavalier-smith, Susanne Menden-deuer, Patrick J. KeelingAbstract:We have sequenced small-subunit (SSU) ribosomal RNA (rRNA) genes from 16 Dinoflagellates, produced phylogenetic trees of the group containing 105 taxa, and combined small- and partial large-subunit (LSU) rRNA data to produce new phylogenetic trees. We compare phylogenetic trees based on Dinoflagellate rRNA and protein genes with established hypotheses of Dinoflagellate evolution based on morphological data. Protein-gene trees have too few species for meaningful in-group phylogenetic analyses, but provide important insights on the phylogenetic position of Dinoflagellates as a whole, on the identity of their close relatives, and on specific questions of evolutionary history. Phylogenetic trees obtained from Dinoflagellate SSU rRNA genes are generally poorly resolved, but include by far the most species and some well-supported clades. Combined analyses of SSU and LSU somewhat improve support for several nodes, but are still weakly resolved. All analyses agree on the placement of Dinoflagellates with ciliates and apicomplexans (=Sporozoa) in a well-supported clade, the alveolates. The closest relatives to dinokaryotic Dinoflagellates appear to be apicomplexans, Perkinsus, Parvilucifera, syndinians and Oxyrrhis. The position of Noctiluca scintillans is unstable, while Blastodiniales as currently circumscribed seems polyphyletic. The same is true for Gymnodiniales: all phylogenetic trees examined (SSU and LSU-based) suggest that thecal plates have been lost repeatedly during Dinoflagellate evolution. It is unclear whether any gymnodinialean clades originated before the theca. Peridiniales appear to be a paraphyletic group from which other Dinoflagellate orders like Prorocentrales, Dinophysiales, most Gymnodiniales, and possibly also Gonyaulacales originated. Dinophysiales and Suessiales are strongly supported holophyletic groups, as is Gonyaulacales, although with more modest support. Prorocentrales is a monophyletic group only in some LSU-based trees. Within Gonyaulacales, molecular data broadly agree with classificatory schemes based on morphology. Implications of this taxonomic scheme for the evolution of selected Dinoflagellate features (the nucleus, mitosis, flagella and photosynthesis) are discussed.
-
multiple protein phylogenies show that oxyrrhis marina and perkinsus marinus are early branches of the Dinoflagellate lineage
International Journal of Systematic and Evolutionary Microbiology, 2003Co-Authors: Juan F. Saldarriaga, Michelle L Mcewan, Naomi M Fast, F J R Taylor, Patrick J. KeelingAbstract:Oxyrrhis marina and Perkinsus marinus are two alveolate species of key taxonomic position with respect to the divergence of apicomplexans and Dinoflagellates. New sequences from Oxyrrhis, Perkinsus and a number of Dinoflagellates were added to datasets of small-subunit (SSU) rRNA, actin, α-tubulin and β-tubulin sequences, as well as to a combined dataset of all three protein-coding genes, and phylogenetic trees were inferred. The parasitic Perkinsus marinus branches at the base of the Dinoflagellate clade with high support in most of the individual gene trees and in the combined analysis, strongly confirming the position originally suggested in previous SSU rRNA and actin phylogenies. The SSU rRNA from Oxyrrhis marina is extremely divergent, and it typically branches with members of the Gonyaulacales, a Dinoflagellate order where SSU rRNA sequences are also divergent. Conversely, none of the three protein-coding genes of Oxyrrhis is noticeably divergent and, in trees based on all three proteins individually and in combination, Oxyrrhis branches at the base of the Dinoflagellate clade, typically with high bootstrap support. In some trees, Oxyrrhis and Perkinsus are sisters, but most analyses indicate that Perkinsus diverged prior to Oxyrrhis. Morphological characters have previously pointed to Oxyrrhis as an early branch in the Dinoflagellate lineage; our data support this suggestion and significantly bolster the molecular data that support a relationship between Perkinsus and Dinoflagellates. Together, these two organisms can be instrumental in reconstructing the early evolution of Dinoflagellates and apicomplexans by helping to reveal aspects of the ancestors of both groups.
-
Dinoflagellate nuclear ssu rrna phylogeny suggests multiple plastid losses and replacements
Journal of Molecular Evolution, 2001Co-Authors: Juan F. Saldarriaga, Patrick J. Keeling, F J R Taylor, Thomas CavaliersmithAbstract:Dinoflagellates are a trophically diverse group of protists with photosynthetic and non-photosynthetic members that appears to incorporate and lose endosymbionts relatively easily. To trace the gain and loss of plastids in Dinoflagellates, we have sequenced the nuclear small subunit rRNA gene of 28 photosynthetic and four non-photosynthetic species, and produced phylogenetic trees with a total of 81 Dinoflagellate sequences. Patterns of plastid gain, loss, and replacement were plotted onto this phylogeny. With the exception of the apparently early-diverging Syndiniales and Noctilucales, all non-photosynthetic Dinoflagellates are very likely to have had photosynthetic ancestors with peridinin-containing plastids. The same is true for all Dinoflagellates with plastids other than the peridinin-containing plastid: their ancestors have replaced one type of plastid for another, in some cases most likely through a non-photosynthetic intermediate. Eight independent instances of plastid loss and three of replacement can be inferred from existing data, but as more non-photosynthetic lineages are characterized these numbers will surely grow.
Juan F. Saldarriaga - One of the best experts on this subject based on the ideXlab platform.
-
Molecular data and the evolutionary history of Dinoflagellates
European Journal of Protistology, 2004Co-Authors: Juan F. Saldarriaga, F. J. R. “max” Taylor, Thomas Cavalier-smith, Susanne Menden-deuer, Patrick J. KeelingAbstract:We have sequenced small-subunit (SSU) ribosomal RNA (rRNA) genes from 16 Dinoflagellates, produced phylogenetic trees of the group containing 105 taxa, and combined small- and partial large-subunit (LSU) rRNA data to produce new phylogenetic trees. We compare phylogenetic trees based on Dinoflagellate rRNA and protein genes with established hypotheses of Dinoflagellate evolution based on morphological data. Protein-gene trees have too few species for meaningful in-group phylogenetic analyses, but provide important insights on the phylogenetic position of Dinoflagellates as a whole, on the identity of their close relatives, and on specific questions of evolutionary history. Phylogenetic trees obtained from Dinoflagellate SSU rRNA genes are generally poorly resolved, but include by far the most species and some well-supported clades. Combined analyses of SSU and LSU somewhat improve support for several nodes, but are still weakly resolved. All analyses agree on the placement of Dinoflagellates with ciliates and apicomplexans (=Sporozoa) in a well-supported clade, the alveolates. The closest relatives to dinokaryotic Dinoflagellates appear to be apicomplexans, Perkinsus, Parvilucifera, syndinians and Oxyrrhis. The position of Noctiluca scintillans is unstable, while Blastodiniales as currently circumscribed seems polyphyletic. The same is true for Gymnodiniales: all phylogenetic trees examined (SSU and LSU-based) suggest that thecal plates have been lost repeatedly during Dinoflagellate evolution. It is unclear whether any gymnodinialean clades originated before the theca. Peridiniales appear to be a paraphyletic group from which other Dinoflagellate orders like Prorocentrales, Dinophysiales, most Gymnodiniales, and possibly also Gonyaulacales originated. Dinophysiales and Suessiales are strongly supported holophyletic groups, as is Gonyaulacales, although with more modest support. Prorocentrales is a monophyletic group only in some LSU-based trees. Within Gonyaulacales, molecular data broadly agree with classificatory schemes based on morphology. Implications of this taxonomic scheme for the evolution of selected Dinoflagellate features (the nucleus, mitosis, flagella and photosynthesis) are discussed.
-
multiple protein phylogenies show that oxyrrhis marina and perkinsus marinus are early branches of the Dinoflagellate lineage
International Journal of Systematic and Evolutionary Microbiology, 2003Co-Authors: Juan F. Saldarriaga, Michelle L Mcewan, Naomi M Fast, F J R Taylor, Patrick J. KeelingAbstract:Oxyrrhis marina and Perkinsus marinus are two alveolate species of key taxonomic position with respect to the divergence of apicomplexans and Dinoflagellates. New sequences from Oxyrrhis, Perkinsus and a number of Dinoflagellates were added to datasets of small-subunit (SSU) rRNA, actin, α-tubulin and β-tubulin sequences, as well as to a combined dataset of all three protein-coding genes, and phylogenetic trees were inferred. The parasitic Perkinsus marinus branches at the base of the Dinoflagellate clade with high support in most of the individual gene trees and in the combined analysis, strongly confirming the position originally suggested in previous SSU rRNA and actin phylogenies. The SSU rRNA from Oxyrrhis marina is extremely divergent, and it typically branches with members of the Gonyaulacales, a Dinoflagellate order where SSU rRNA sequences are also divergent. Conversely, none of the three protein-coding genes of Oxyrrhis is noticeably divergent and, in trees based on all three proteins individually and in combination, Oxyrrhis branches at the base of the Dinoflagellate clade, typically with high bootstrap support. In some trees, Oxyrrhis and Perkinsus are sisters, but most analyses indicate that Perkinsus diverged prior to Oxyrrhis. Morphological characters have previously pointed to Oxyrrhis as an early branch in the Dinoflagellate lineage; our data support this suggestion and significantly bolster the molecular data that support a relationship between Perkinsus and Dinoflagellates. Together, these two organisms can be instrumental in reconstructing the early evolution of Dinoflagellates and apicomplexans by helping to reveal aspects of the ancestors of both groups.
-
Dinoflagellate nuclear ssu rrna phylogeny suggests multiple plastid losses and replacements
Journal of Molecular Evolution, 2001Co-Authors: Juan F. Saldarriaga, Patrick J. Keeling, F J R Taylor, Thomas CavaliersmithAbstract:Dinoflagellates are a trophically diverse group of protists with photosynthetic and non-photosynthetic members that appears to incorporate and lose endosymbionts relatively easily. To trace the gain and loss of plastids in Dinoflagellates, we have sequenced the nuclear small subunit rRNA gene of 28 photosynthetic and four non-photosynthetic species, and produced phylogenetic trees with a total of 81 Dinoflagellate sequences. Patterns of plastid gain, loss, and replacement were plotted onto this phylogeny. With the exception of the apparently early-diverging Syndiniales and Noctilucales, all non-photosynthetic Dinoflagellates are very likely to have had photosynthetic ancestors with peridinin-containing plastids. The same is true for all Dinoflagellates with plastids other than the peridinin-containing plastid: their ancestors have replaced one type of plastid for another, in some cases most likely through a non-photosynthetic intermediate. Eight independent instances of plastid loss and three of replacement can be inferred from existing data, but as more non-photosynthetic lineages are characterized these numbers will surely grow.
-
Dinoflagellate phylogeny revisited: reconciling morphological and molecular based phylogenies
Grana, 1999Co-Authors: Robert A. Fensome, Juan F. Saldarriaga, “max” F. J. R. TaylorAbstract:Ultrastructural and molecular phylogenetic data suggest that Dinoflagellates diverged as a lineage possibly as early as the Precambrian. However, the fossil record is problematic before the Mesozoic. From the mid Triassic, though, the fossil record of Dinoflagellates is a rich source of information on Mesozoic-Cenozoic Dinoflagellates, especially the gonyaulacoids and peridinioids. From the sequence of appearance of species and tabulation types and the impression of early morphological experimentation and later stabilization, the early Mesozoic radiation of Dinoflagellates appears to be a real evolutionary event: indeed, Dinoflagellate morphology as we know it today may originate in that event. This would explain why it is so difficult to interpret earlier fossils as Dinoflagellates. However, that the Dinoflagellate lineage existed in some form in the pre-Mesozoic is supported by biogeochemical data, early results of which indicate that certain early Paleozoic acanthomorph acritarchs may belong to the lin...
