240 research outputs found
Phylogenomic Insights into the Origin of Primary Plastids
No full textAbstract The origin of plastids was a major evolutionary event that paved the way for an astonishing diversification of photosynthetic eukaryotes. Plastids originated by endosymbiosis between a heterotrophic eukaryotic host and cyanobacteria, presumably in a common ancestor of the primary photosynthetic eukaryotes (Archaeplastida). A single origin of primary plastids is well supported by plastid evidence but not by nuclear phylogenomic analyses, which have consistently failed to recover the monophyly of Archaeplastida hosts. Importantly, plastid monophyly and nonmonophyletic hosts could be explained under scenarios of independent or serial eukaryote-to-eukaryote endosymbioses. Here, we assessed the strength of the signal for the monophyly of Archaeplastida hosts in four available phylogenomic data sets. The effect of phylogenetic methodology, data quality, alignment trimming strategy, gene and taxon sampling, and the presence of outlier genes were investigated. Our analyses revealed a lack of support for host monophyly in the shorter individual data sets. However, when analyzed together under rigorous data curation and complex mixture models, the combined nuclear data sets supported the monophyly of primary photosynthetic eukaryotes (Archaeplastida) and recovered a putative association with plastid-lacking Picozoa. This study represents an important step toward better understanding deep eukaryotic evolution and the origin of plastids. [Archaeplastida; Bayesian; chloroplast; maximum likelihood; mixture model; ortholog; outlier loci; paralog; protist.
Author Correction: A molecular timescale for eukaryote evolution with implications for the origin of red algal-derived plastids
No full textA molecular timescale for eukaryote evolution with implications for the origin of red algal-derived plastids
No full textIn modern oceans, eukaryotic phytoplankton is dominated by lineages with red algal-derived plastids such as diatoms, dinoflagellates, and coccolithophores. Despite the ecological importance of these groups and many others representing a huge diversity of forms and lifestyles, we still lack a comprehensive understanding of their evolution and how they obtained their plastids. New hypotheses have emerged to explain the acquisition of red algal-derived plastids by serial endosymbiosis, but the chronology of these putative independent plastid acquisitions remains untested. Here, we establish a timeframe for the origin of red algal-derived plastids under scenarios of serial endosymbiosis, using Bayesian molecular clock analyses applied on a phylogenomic dataset with broad sampling of eukaryote diversity. We find that the hypotheses of serial endosymbiosis are chronologically possible, as the stem lineages of all red plastid-containing groups overlap in time. This period in the Meso- and Neoproterozoic Eras set the stage for the later expansion to dominance of red algal-derived primary production in the contemporary oceans, which profoundly altered the global geochemical and ecological conditions of the Earth
eine Analyse von Ultrastruktur, Phylogenie und Kospeziation
No full textThe complex mechanisms leading to a tripartite symbiosis involving bacteria,
flagellates, and host termites are not yet fully understood. While the
flagellates are known to play a major role in the degradation of the
cellulosic food of the termites, in most cases, the functions of the diverse
flagellate-associated bacteria are completely obscure. Unambiguous
identification of the mostly uncultivable prokaryotes and eukaryotes is an
important step in understanding the mutual interactions between the two
partners. For this purpose, in the studies described in my thesis,
morphological investigations (light microscopy and electron microscopy) were
combined with molecular phylogenetic analyses (full-cycle-rRNA approach). In
two earlier light microscopy studies, other authors reported contradicting
numbers of devescovinid flagellates occurring in the hindgut of the dry-wood
termite Incisitermes marginipennis. We clearly and unambiguously documented
the presence of only one devescovinid species (Metadevescovina modica)
inhabiting the gut of I. marginipennis using a combination of various light
and electron microscopy techniques and molecular phylogenetic analysis of the
small subunit (SSU) rRNA gene sequences. Moreover, we confirmed the validity
of the genus Metadevescovina, which had long been discussed as being the same
as the genus Devescovina; monophyly of each of the genera was revealed by
molecular phylogenetic analyses. Metadevescovina could not be distinguished
from Devescovina solely by morphological characteristics of the flagellates
themselves, but the two flagellate genera could be differentiated by examining
their bacterial symbionts. The cell surface of Metadevescovina flagellates is
densely colonized with spirochetes, and that of Devescovina flagellates is
densely covered with filamentous bacteria affiliated to the Bacteroidales.
Molecular phylogenetic analyses of Devescovina spp. and their bacterial
symbionts from a wide range of Kalotermitidae revealed that the termites
acquired two bacterial symbionts by two different routes: vertical
transmission and horizontal transmission. The ectosymbionts of Devescovina
spp. form a monophyletic group within the Bacteroidales (“Candidatus
Armantifilum devescovinae”). Congruence analyses of the phylogenetic trees of
Devescovina spp. and “Candidatus Armantifilum devescovinae” documented a
strict cospeciation of the partners, which indicated an obligate symbiosis,
leading to a vertical transmission of the bacteria within their host lineages.
The ‘Endomicrobia’ endosymbionts of Devescovina spp. are most closely related
to endosymbionts of phylogenetically unrelated termite gut flagellates, which
indicated that these symbionts were acquired by horizontal transmission
between different flagellate species present in the same termite gut. In a
further study documented in this thesis, the multiple symbionts of the
flagellate Joenia annectens from the dry-wood termite Kalotermes flavicollis
were identified, localized using a full-cycle-rRNA approach, and
morphologically described at the ultrastructural level. Two populations of J.
annectens could be distinguished not only by their SSU rRNA gene sequences
(0.8% sequence divergence), but also by differences in their assemblages of
bacterial symbionts. Each of the flagellate populations hosted
phylogenetically distinct ectosymbionts from the phylum Bacteroidetes, while a
single phylotype of ‘Endomicrobia’ was consistently associated with only one
of the host phylotypes. However, not all individuals were colonized, once
again corroborating that ‘Endomicrobia’ are not always cospeciating with their
host lineages. The results reported in my thesis provide important information
about the specificity of the symbioses between termite gut flagellates and
their bacterial symbionts. This information is necessary for further studies
of the function of these symbioses. A possible involvement of bacterial
symbionts in the nitrogen metabolism of the host flagellates is discussed.Die komplexen Mechanismen, welche der Dreiersymbiose zwischen Bakterien,
Flagellaten und ihren Wirtstermiten zugrundeliegen, sind bis heute noch
weitgehend unverstanden. Während den Flagellaten eine wesentliche Rolle beim
Abbau der cellulosehaltigen Nahrung der Termiten zugeschrieben wird, sind die
Funktionen der diversen, mit den Flagellaten assoziierten Bakterien in den
meisten Fällen komplett unbekannt. Ein wichtiger Schritt für die Erforschung
der wechselseitigen Beziehungen zwischen den in der Regel nicht kultivierbaren
pro- und eukaryotischen Symbionten stellt deren eindeutige Identifizierung
dar. Dazu wurden in den vorliegenden Studien morphologische Untersuchungen
(Licht- und Elektronenmikroskopie) beider Partner mit molekular-
phylogenetischen Analysen kombiniert (full-cycle-rRNA approach). In zwei
vorangegangenen lichtmikroskopischen Studien anderer Autoren wurde für die
Trockenholztermite Incisitermes marginipennis eine widersprüchliche
Artenanzahl von devescoviniden Flagellaten dokumentiert. Durch den Einsatz
verschiedener licht- und elektronenmikroskopischer Techniken sowie durch die
Analyse der Sequenzvariabilität der Gene der kleinen ribosomalen Untereinheit
(SSU rRNA) konnten wir in der vorliegenden Arbeit eindeutig zeigen, dass I.
marginipennis lediglich eine Art devescovinider Flagellaten (Metadevescovina
modica) im Darm beherbergt. Gleichzeitig konnten wir die Gültigkeit der
Gattung Metadevescovina, welche bis zum heutigen Tag stark umstritten war und
häufig als Synonym zu Devescovina angesehen wurde, bestätigen.
Molekularphylogenetische Analysen zeigten, dass beide Gattungen jeweils eine
separate monophyletische Gruppe bilden. Eine Unterscheidung der beiden
Gattungen an Hand morphologischer Merkmale der Flagellaten selbst war nicht
möglich, konnte jedoch unter Berücksichtigung ihrer bakteriellen Symbionten
erfolgen. Während Flagellaten der Gattung Metadevescovina einen dichten Besatz
von Spirochaeten auf ihrer Oberfläche zeigen, sind Devescovina spp.
vollständig von filamentösen Bakterien bedeckt, welche den Bacteroidales
zugeordnet werden. Molekularphylogenetische Analysen von Devescovina spp. von
verschiedenen Vertretern der Kalotermitidae und ihren bakteriellen Symbionten
ergaben zwei verschiedene Szenarien bezüglich des Erwerbs dieser Symbionten:
Eine vertikale Weitergabe und eine horizontale Weitergabe. Es konnte gezeigt
werden, dass die Ektosymbionten eine monophyletische Gruppe innerhalb der
Bacteroidales bilden („Candidatus Armantifilum devescovinae“).
Kongruenzanalysen der Stammbäume von Devescovina spp. und „Candidatus
Armantifilum devescovinae“ dokumentierten eine strikte Kospeziation der
Partner. Eine obligate Symbiose der beiden Partner, und somit eine vertikale
Weitergabe der Bakterien innerhalb ihrer Wirtsflagellaten, konnte demnach
belegt werden. Der Erwerb von Symbionten durch horizontale Weitergabe von
anderen Wirtsflagellaten wurde dagegen für die im Zytoplasma vorkommenden
‚Endomicrobia‘ dokumentiert. Hier waren die nächsten Verwandten der mit den
Devescovina spp. assoziierten ‚Endomicrobia‘ Endosymbionten von phylogenetisch
nicht verwandten Termitenflagellaten. In einer weiteren Studie dieser Arbeit
wurden die multiplen Symbionten des Flagellaten Joenia annectens aus der
Trockenholztermite Kalotermes flavicollis identifiziert und lokalisiert (full-
cycle-rRNA approach). Ultrastrukturelle Untersuchungen ermöglichten eine
morphologische Beschreibung der gefundenen Phylotypen. Basierend auf den
Assoziationen mit phylogenetisch verschiedenen Symbionten konnten zwei
Populationen von J. annectens unterschieden werden. Gestützt wurde das
Ergebnis durch die Analyse der SSU rRNA Gensequenzen von J. annectens (0,8%
Sequenzunterschied zwischen beiden Populationen). Beide
Flagellatenpopulationen waren jeweils mit eigenen Ektosymbionten des Phylums
Bacteroidetes assoziiert. Dahingegen beherbergte nur eine der beiden
Populationen von J. annectens einen Vertreter der ‚Endomicrobia‘ im
Zytoplasma. Das Fehlen von ‚Endomicrobia‘-Symbionten bei vielen Flagellaten
der gleichen Population zeigt ein weiteres Beispiel dafür, dass diese
Symbionten nicht immer mit ihren Wirtsflagellaten kospeziieren. Die Ergebnisse
meiner Arbeit haben wichtige Erkenntnisse zur Spezifität der Symbiosen
zwischen Termitenflagellaten und ihrer bakteriellen Symbionten gebracht. Sie
stellen somit eine Grundvoraussetzung für die anstehende Erforschung der
funktionellen Aspekte dieser Symbiosen dar. Eine Beteiligung der bakteriellen
Symbionten am Stickstoffstoffwechsel der Flagellaten wird diskutiert
Early Diversification of Membrane Intrinsic Proteins (MIPs) in Eukaryotes
No full textAbstract Membrane intrinsic proteins (MIPs), including aquaporins (AQPs) and aquaglyceroporins (GLPs), form an ancient family of transporters for water and small solutes across biological membranes. The evolutionary history and functions of MIPs have been extensively studied in vertebrates and land plants, but their widespread presence across the eukaryotic tree of life suggests both a more complex evolutionary history and a broader set of functions than previously thought. That said, the early evolution of MIPs remains obscure. The presence of one GLP and four AQP clades across both bacteria and archaea suggests that the first eukaryotes could have possessed up to five MIPs. Here, we report on a previously unknown richness in MIP diversity across all major eukaryotic lineages, including unicellular eukaryotes, which make up the bulk of eukaryotic diversity. Three MIP clades have likely deep evolutionary origins, dating back to the last eukaryotic common ancestor (LECA), and support the presence of a complex MIP repertoire in early eukaryotes. Overall, our findings highlight the growing complexity of the reconstructed LECA genome: the dynamic evolutionary history of MIPs was set in motion when eukaryotes were in their infancy followed by radiative bursts across all main eukaryotic lineages.Open-Access-Publikationsfonds 202
Predatory colponemids are the sister group to all other alveolates
No full textAlveolates are a major supergroup of eukaryotes encompassing more than ten thousand free-living and parasitic species, including medically, ecologically, and economically important apicomplexans, dinoflagellates, and ciliates. These three groups are among the most widespread eukaryotes on Earth, and their environmental success can be linked to unique innovations that emerged early in each group. Understanding the emergence of these well-studied and diverse groups and their innovations has relied heavily on the discovery and characterization of early-branching relatives, which allow ancestral states to be inferred with much greater confidence. Here we report the phylogenomic analyses of 313 eukaryote protein-coding genes from transcriptomes of three members of one such group, the colponemids (Colponemidia), which support their monophyly and position as the sister lineage to all other known alveolates. Colponemid-related sequences from environmental surveys and our microscopical observations show that colponemids are not common in nature, but they are diverse and widespread in freshwater habitats around the world. Studied colponemids possess two types of extrusive organelles (trichocysts or toxicysts) for active hunting of other unicellular eukaryotes and potentially play an important role in microbial food webs. Colponemids have generally plesiomorphic morphology and illustrate the ancestral state of Alveolata. We further discuss their importance in understanding the evolution of alveolates and the origin of myzocytosis and plastids
Evaluation of the hCMEC/D3 cell line, a new "in vitro" model of the human blood-brain barrier for transport and gene regulation studies
No full textBrain endothelial capillary cells form the blood-brain barrier (BBB), a highly selective
membrane between the peripheral blood and the central nervous system. The main
functions of the BBB are to protect the brain tissue by preventing the entry of toxic
compounds and to supply it with nutrients in order to assure proper function. Tight
junctions are the key elements for the establishment of a tight barrier and seal the
intercellular gaps against passive diffusion of hydrophilic compounds. A second important
characteristic of the brain capillary endothelial cells are transport proteins that prevent
brain penetration of their substrates by pumping them back in the blood. These
compounds include a series of clinically used drugs. Important drug efflux transporters
located at the BBB are P-glycoprotein (P-gp), the breast cancer resistance protein
(BCRP) and the family of multidrug resistance proteins (MRP).
During drug development, the question of whether a drug candidate reaches the brain
tissue is of great importance. Therefore, models are needed to predict the BBB
permeability of new compounds. In the past, in vitro models have been developed to
address this question. These models include isolated brain capillaries, isolated primary
brain capillary endothelial cells and BBB cell lines of various origins. A major problem
encountered with these cell lines was an insufficient paracellular resistance.
Recently, the hCMEC/D3 cell line was generated by immortalizing primary human brain
endothelial cells. In culture this cell line shows a morphology that closely resembles to
primary cells, forms tight monolayers and expresses BBB markers such as chemokine
receptors, tight junctional molecules and ATP binding cassette (ABC)-transporters.
The aim of this thesis was to evaluate the hCMEC/D3 cell line as an in vitro model of the
human BBB to study 1) permeability properties including para- and transcellular diffusion
as well as active transport 2) the influence of endo- and exogenous factors on the
paracellular permeability and 3) the regulation of breast cancer resistance protein and Pglycoprotein
by pro-inflammatory cytokines.
The first study describes the characterization of the hCMEC/D3 cells as an in vitro model
of the human BBB for permeability studies (section Error! Reference source not
found.). The ability of the cells to allow discrimination between para- and transcellular
diffusion was investigated by measuring the transport of a series of compounds with
different physicochemical properties. A ratio of 2.8 was observed when comparing the
permeabilities of the compounds with the highest and the lowest diffusion rate. The
passive permeability of sucrose could be reduced significantly by replacing fetal calf
serum with human serum. Furthermore, quantitative mRNA expression of the ABCtransporters
P-gp, BCRP, MRP1, MRP2, MRP3, MRP4, MRP5 as well as the human
transferrin receptor (hTfR) was shown. Protein expression of P-gp, BCRP and the hTfR
was detected and functional activity of P-gp, BCRP and the MRPs was investigated in
efflux experiments. Furthermore, bidirectional P-gp transport activity was observed.
In a second project the impact of endo- and exogenous factors on the paracellular
permeability of hCMEC/D3 monolayers was assessed, since it is know that the molecular
assembly of tight junctions depends on the surrounding milieu (section Error! Reference
source not found.). Based on reports in the literature, the cells were incubated with a
variety of compounds that included anti-inflammatory drugs, growth factors and
antioxidants. The effects on the monolayer tightness of hCMEC/D3 were investigated by
measuring the transport of sucrose, a paracellular permeability marker. N-acetylcystein
(NAC), atorvastatin and sodium nitroprusside (SNP) reduced the sucrose permeability
significantly, and slightly increased zonula occludens protein (ZO-1) expression.
Additionally, NAC and SNP reduced the generation of reactive oxygen species (ROS),
which have been reported to disrupt the assembly of tight junctions.
The effect of the pro-inflammatory cytokines IL-1[beta], IL-6 and TNF-[alpha] on the expression and
activity of the ABC-transporters BCRP and P-gp was investigated in the hCMEC/D3 cell
line (section Error! Reference source not found.). IL-1[beta], IL-6 and TNF-[alpha], which are
know to be elevated during various diseases, suppressed significantly BCRP mRNA
expression. In addition, BCRP activity was reduced under the influence of all tested
cytokines, as shown by efflux experiments. P-gp mRNA levels were slightly reduced by
IL-6 but significantly increased after TNF-[alpha] treatment. TNF-[alpha] also increased the protein
expression of P-gp. This in vitro study indicates that expression levels of BCRP and P-gp
at the BBB might be altered during acute or chronic inflammation, resulting in a changed
brain penetration of their substrates.
In an isolated project, the pharmacokinetics and pharmacodynamics of increasing oral
doses of the satiety peptides GLP-1 and PYY3-36 were assessed in healthy male
volunteers. Oral administration of either peptide induced a rapid and dose-dependent
increase in plasma drug concentrations. Oral administration of GLP-1 induced a potent
effect on insulin release and both peptides suppressed ghrelin secretion. In conclusion,
this study showed, for the first time, that satiety peptides such as GLP-1 and PYY3-36
can be orally delivered safely and effectively in humans
Freedom and the 'creative act' in the writings of Nikolai Berdiaev : an evaluation in light of Jürgen Moltmann's theology of freedom
No full textThis project revisits the work of Nikolai Berdiaev, one of the first Russian Silver Age religious philosophers to be widely read in the West. The focus of this research is his thought on freedom and the ‘creative act’. We will argue that Berdiaev’s vision of freedom contains two types of freedom – a freedom understood within the created order and a freedom ‘outside’ of creation. It will be shown that in the former type, the reader finds a nuanced and insightful multi-layered conception of human freedom, which offers intriguing possibilities for exploring freedom and its implications for humanity. It will also be demonstrated that this type of freedom is closely related to his innovative view of creativity. Berdiaev conceives of freedom and creativity as distinct concepts, and yet so integrally related that they are interdependent. In the latter type of freedom, the reader will encounter a highly speculative and original metaphysical view that attempts to explain freedom as non-determination and answer the challenges of theodicy, which, this research will maintain, fails to do.
This research will contend (contrary to Berdiaev’s own statements) that his thought is most comprehensible from a broadly theological perspective. This perspective will underscore the significant tension within his work that arises from his speculative metaphysics. Unlike earlier works on Berdiaev that glossed over this tension, we will attempt to ameliorate it by engaging Jürgen Moltmann’s theology of freedom. Moltmann’s theology will provide a number of ideas and concepts for an analysis, critique, and reconfiguration of Berdiaev’s vision. This reconfiguration will seek to remain faithful to Berdiaev’s core concerns, while providing a new interpretation of his thought that is relevant for a contemporary dialogue concerning the significance of freedom and creativity for the person and community in relation to God
Single cell genomics of uncultured marine alveolates shows paraphyly of basal dinoflagellates
No full textMarine alveolates (MALVs) are diverse and widespread early-branching dinoflagellates, but most knowledge of the group comes from a few cultured species that are generally not abundant in natural samples, or from diversity analyses of PCR-based environmental SSU rRNA gene sequences. To more broadly examine MALV genomes, we generated single cell genome sequences from seven individually isolated cells. Genes expected of heterotrophic eukaryotes were found, with interesting exceptions like presence of proteorhodopsin and vacuolar H+-pyrophosphatase. Phylogenetic analysis of concatenated SSU and LSU rRNA gene sequences provided strong support for the paraphyly of MALV lineages. Dinoflagellate viral nucleoproteins were found only in MALV groups that branched as sister to dinokaryotes. Our findings indicate that multiple independent origins of several characteristics early in dinoflagellate evolution, such as a parasitic life style, underlie the environmental diversity of MALVs, and suggest they have more varied trophic modes than previously thought
Phylogenomic Insights into the Origin of Primary Plastids
No full textThe origin of plastids was a major evolutionary event that paved the way for an astonishing diversification of photosynthetic eukaryotes. Plastids originated by endosymbiosis between a heterotrophic eukaryotic host and cyanobacteria, presumably in a common ancestor of the primary photosynthetic eukaryotes (Archaeplastida). A single origin of primary plastids is well supported by plastid evidence but not by nuclear phylogenomic analyses, which have consistently failed to recover the monophyly of Archaeplastida hosts. Importantly, plastid monophyly and nonmonophyletic hosts could be explained under scenarios of independent or serial eukaryote-to-eukaryote endosymbioses. Here, we assessed the strength of the signal for the monophyly of Archaeplastida hosts in four available phylogenomic data sets. The effect of phylogenetic methodology, data quality, alignment trimming strategy, gene and taxon sampling, and the presence of outlier genes were investigated. Our analyses revealed a lack of support for host monophyly in the shorter individual data sets. However, when analyzed together under rigorous data curation and complex mixture models, the combined nuclear data sets supported the monophyly of primary photosynthetic eukaryotes (Archaeplastida) and recovered a putative association with plastid-lacking Picozoa. This study represents an important step toward better understanding deep eukaryotic evolution and the origin of plastids.</p
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