22333
2021
eng
doctoralthesis
1
2021-02-06
--
2021-02-05
Expansion Microscopy (ExM) as a tool to study organelles and intracellular pathogens
Expansionsmikroskopie (ExM) als Tool zur Untersuchung von Organellen und intrazellulären Pathogenen
The resolution of fluorescence light microscopy was long believed to be limited by the diffraction limit of light of around 200-250 nm described in 1873 by Ernst Abbe. Within the last decade, several approaches, such as structured illumination microscopy (SIM), stimulated emission depletion STED and (direct) stochastic optical reconstruction microscopy (d)STORM have been established to bypass the diffraction limit. However, such super-resolution techniques enabling a resolution <100 nm require specialized and expensive setups as well as expert knowledge in order to avoid artifacts. They are therefore limited to specialized laboratories. Recently, Boyden and colleagues introduced an alternate approach, termed expansion microscopy (ExM). The latter offers the possibility to perform superresolution microscopy on conventional confocal microscopes by embedding the sample into a swellable hydrogel that is isotropically expanded. Since its introduction in 2015, expansion microscopy has developed rapidly offering protocols for 4x, 10x and 20x expansion of proteins and RNA in cells, tissues and human clinical specimens.
Mitochondria are double membrane-bound organelles and crucial to the cell by performing numerous tasks, from ATP production through oxidative phosphorylation, production of many important metabolites, cell signaling to the regulation of apoptosis. The inner mitochondrial membrane is strongly folded forming so-called cristae. Besides being the location of the oxidative phosphorylation and therefore energy conversion and ATP production, cristae have been of great interest because changes in morphology have been linked to a plethora of diseases from cancer, diabetes, neurodegenerative diseases, to aging and infection. However, cristae imaging remains challenging as the distance between two individual cristae is often below 100 nm. Within this work, we demonstrate that the mitochondrial creatine kinase MtCK linked to fluorescent protein GFP (MtCK-GFP) can be used as a cristae marker. Upon fourfold expansion, we illustrate that our novel marker enables visualization of cristae morphology and localization of mitochondrial proteins relative to cristae without the need for specialized setups. Furthermore, we show the applicability of expansion microscopy for several bacterial pathogens, such as Chlamydia trachomatis, Simkania negevensis, Neisseria gonorrhoeae and Staphylococcus aureus. Due to differences in bacterial cell walls, we reveal important aspects for the digestion of pathogens for isotropic expansion. We further show that expansion of the intracellular pathogens C. trachomatis and S. negevensis, enables the differentiation between the two distinct developmental forms, catabolic active reticulate bodies (RB) and infectious elementary bodies (EB), on a conventional confocal microscope. We demonstrate the possibility to precisely locate chlamydial effector proteins, such as CPAF or Cdu1, within and outside the chlamydial inclusion. Moreover, we show that expansion microscopy enables the investigation of bacteria, herein S. aureus, within LAMP1 and LC3-II vesicles. With the introduction of the unnatural α-NH2-ω-N3-C6-ceramide, we further present the first approach for the expansion of lipids that may also be suitable for far inaccessible molecule classes like carbohydrates. The efficient accumulation and high labeling density of our functionalized α-NH2-ω-N3-C6-ceramide in both cells and bacteria enables in combination with tenfold expansion nanoscale resolution (10-20 nm) of the interaction of proteins with the plasma membrane, membrane of organelles and bacteria. Ceramide is the central molecule of the sphingolipid metabolism, an important constituent of cellular membranes and regulates many important cellular processes such as differentiation, proliferation and apoptosis. Many studies report about the importance of sphingolipids during infection of various pathogens. While the transport of ceramide to Chlamydia has been reported earlier, one of the unanswered questions remaining was if ceramide forms parts of the outer or inner bacterial membrane. Expansion of α-NH2-ω-N3-C6-ceramide enabled the visualization of ceramide in the inner and outer membrane of C. trachomatis and their distance was determined to be 27.6 ± 7.7 nm.
Aufgrund der Beugungseigenschaften des Lichtes wurde bereits 1873 durch Ernst Abbe für die Lichtmikroskopie eine theoretische Auflösungsgrenze von 200-250 nm definiert. Durch die Einführung verschiedener hochauflösender Mikroskopiemethoden, wie beispielsweise SIM-Mikroskopie (structured illumination microscopy), STED-Mikroskopie (stimulated emission depletion) und (d)STORM-Mikroskopie ((direct) stochastic optical reconstruction microscopy), konnte im letzten Jahrzehnt jedoch die Auflösung auf unter 100 nm verbessert werden. Allerdings benötigen solche Hochauflösungstechniken sowohl spezialisierte und kostenintensive Geräte als auch Expertenwissen zur Vermeidung von Artefakten, sodass diese nur in wenigen Laboren angewendet werden können. Ein alternativer Ansatz, die sogenannte Expansionsmikroskopie, wurde kürzlich von der Arbeitsgruppe um Ed Boyden etabliert. Hierbei wird eine Probe mit einem quellfähigen Gel vernetzt, welches daraufhin isotrop expandiert wird, sodass auch an konventionellen konfokalen Mikroskopen Hochauflösung ermöglicht wird. Seit ihrer Einführung im Jahre 2015 hat sich die Expansionsmikroskopie schnell entwickelt und bietet Protokolle für 4-fache, 10-fache oder sogar 20-fache Expansion von Proteinen als auch RNA in Zellen oder sogar komplexen Geweben.
Mitochondrien besitzen zwei Membranen und sind für die Zelle von großer Bedeutung, da sie eine Vielzahl wichtiger Aufgaben übernehmen - von der ATP-Produktion durch die oxidative Phosphorylierung über die Produktion vieler wichtiger Metabolite bis hin zur Regulation zellulärer Signalwege. Die innere Mitochondrienmembran ist stark gefaltet und bildet Einstülpungen, die sogenannten Cristae, in welchen die oxidative Phosphorylierung und somit die Energieumwandlung und ATP-Synthese stattfindet. Morphologische Veränderungen der Cristae können sowohl beim Altern von Zellen, als auch bei verschiedenen Infektionen beobachtet werden und können darüber hinaus auch im Rahmen diverser Erkrankungen, wie beispielsweise Krebs, Diabetes oder neurodegenerativen Erkrankungen auftreten. Die Visualisierung der Cristae durch Fluoreszenzmikroskopie ist herausfordernd, da der Abstand zwischen einzelnen Cristae oftmals unter 100 nm beträgt. In der vorliegenden Arbeit wird gezeigt, dass die Expression der mitochondrialen Kreatinkinase gekoppelt an das Fluoreszenzprotein GFP (MtCK-GFP) als Cristaemarker genutzt werden kann. In Kombination mit vierfacher Expansion ermöglicht unser Marker die Untersuchung morphologischer Veränderungen von Cristae, sowie die Lokalisierung mitochondrialer Proteine relativ zu den Cristae. Darüber hinaus wird im Rahmen dieser Arbeit die Anwendbarkeit der Expansionsmikroskopie für mehrere bakterielle Pathogene, und zwar Chlamydia trachomatis, Simkania negevensis, Neisseria gonorrhoeae und Staphylococcus aureus, gezeigt. Hierbei verdeutlichen wir wichtige Aspekte für den vollständigen Verdau unterschiedlicher bakterieller Zellwände und somit isotropen Expansion. Die Expansion der intrazellulären Pathogene C. trachomatis und S. negevensis ermöglichte es uns an konventionellen konfokalen Mikroskopen zwischen den zwei verschiedenen Entwicklungsstadien, der katabolisch aktiven Retikulärkörperchen (RBs) und der infektiösen Elementarkörperchen (EBs), zu unterscheiden. Außerdem konnte die Möglichkeit der präzisen Lokalisierung chlamydialer Proteine wie CPAF und Cdu1 innerhalb und außerhalb der chlamydialen Inklusion gezeigt werden und Bakterien, in diesem Fall S. aureus, in LAMP1 und LC3-II Vesikeln visualisiert werden. Mit der Einführung des unnatürlichen α-NH2-ω-N3-C6-Ceramides, präsentieren wir zudem ein erstes Konzept für die Expansion von Lipiden, welches möglicherweise auch für deutlich unzugänglichere Molekülklassen wie beispielsweise Kohlehydrate geeignet ist. Die effiziente Akkumulierung unseres funktionalisierten α-NH2-ω-N3-C6-Ceramides in Zellen sowie Bakterien ermöglicht in Kombination mit zehnfacher Expansion die Untersuchung der Interaktion von Proteinen mit der Zellmembran, Membranen von Organellen und Bakterien mit einer räumlichen Auflösung von 10-20 nm. Ceramid ist das zentrale Molekül des Sphingolipidstoffwechsels, ein wichtiger Baustein zellulärer Membrane und reguliert viele essentielle Prozesse wie die Zelldifferenzierung, die Proliferation als auch die Apoptose. Viele Studien berichten von der Bedeutung der Sphingolipide während der Infektion verschiedener Pathogene. So wurde beispielsweise zuvor berichtet, dass Ceramide aktiv zu Chlamydien transportiert und in deren Membranen eingebaut werden. Hierbei verblieb allerdings die Frage, ob Ceramide in der äußeren oder inneren bakteriellen Membran lokalisiert sind. Die Expansion unseres α-NH2-ω-N3-C6-Ceramides ermöglichte es uns Ceramide in der inneren und äußeren Membran von C. trachomatis zu visualisieren und den Abstand zwischen beiden Membranen auf 27.6 ± 7.7 nm zu bestimmen.
urn:nbn:de:bvb:20-opus-223330
10.25972/OPUS-22333
publish
X 129180
Deutsches Urheberrecht
Tobias C. Kunz
deu
swd
Fluoreszenzmikroskopie
deu
swd
Mitochondrium
deu
swd
Chlamydia trachomatis
deu
swd
Ceramide
eng
uncontrolled
Superresolution microscopy
eng
uncontrolled
Expansion microscopy
eng
uncontrolled
Mitochondria
eng
uncontrolled
Intracellular bacteria
deu
uncontrolled
Hochauflösungsmikroskopie
deu
uncontrolled
Expansionsmikroskopie
deu
uncontrolled
Mitochondrien
deu
uncontrolled
Intrazelluläre Bakterien
Biowissenschaften; Biologie
open_access
Fakultät für Biologie
Graduate School of Life Sciences
Universität Würzburg
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/22333/Kunz_Tobias-Christopher_Dissertation.pdf
23124
2020
eng
11
article
1
2021-03-23
--
--
Nanoscale imaging of bacterial infections by sphingolipid expansion microscopy
Expansion microscopy (ExM) enables super-resolution imaging of proteins and nucleic acids on conventional microscopes. However, imaging of details of the organization of lipid bilayers by light microscopy remains challenging. We introduce an unnatural short-chain azide- and amino-modified sphingolipid ceramide, which upon incorporation into membranes can be labeled by click chemistry and linked into hydrogels, followed by 4x to 10x expansion. Confocal and structured illumination microscopy (SIM) enable imaging of sphingolipids and their interactions with proteins in the plasma membrane and membrane of intracellular organelles with a spatial resolution of 10-20nm. As our functionalized sphingolipids accumulate efficiently in pathogens, we use sphingolipid ExM to investigate bacterial infections of human HeLa229 cells by Neisseria gonorrhoeae, Chlamydia trachomatis and Simkania negevensis with a resolution so far only provided by electron microscopy. In particular, sphingolipid ExM allows us to visualize the inner and outer membrane of intracellular bacteria and determine their distance to 27.6 +/- 7.7nm. Imaging of lipid bilayers using light microscopy is challenging. Here the authors label cells using a short chain click-compatible ceramide to visualize mammalian and bacterial membranes with expansion microscopy.
Nature Communications
10.1038/s41467-020-19897-1
urn:nbn:de:bvb:20-opus-231248
publish
Nature Communications (2020) 11:6173. https://doi.org/10.1038/s41467-020-19897-1
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Ralph Götz
Tobias C. Kunz
Julian Fink
Franziska Solger
Jan Schlegel
Jürgen Seibel
Vera Kozjak-Pavlovic
Thomas Rudel
Markus Sauer
eng
uncontrolled
nanoscale imaging
eng
uncontrolled
bacterial infection
eng
uncontrolled
sphingolipid expansion microscopy
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Institut für Organische Chemie
Förderzeitraum 2020
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/23124/s41467-020-19897-1.pdf
24484
2021
eng
12
article
1
--
2021-07-29
--
Staphylococcus aureus α-Toxin Induces Acid Sphingomyelinase Release From a Human Endothelial Cell Line
Staphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (α-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal α-toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus α-toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal α-toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.
Frontiers in Microbiology
1664-302X
10.3389/fmicb.2021.694489
urn:nbn:de:bvb:20-opus-244843
2021-09-13T09:05:03+00:00
sword
swordwue
attachment; filename=deposit.zip
81784b3002a2d94c9f23295f181cfcd9
Frontiers in Microbiology (2021) 12:694489. doi: 10.3389/fmicb.2021.694489
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
David Krones
Marcel Rühling
Katrin Anne Becker
Tobias C. Kunz
Carolin Sehl
Kerstin Paprotka
Erich Gulbins
Martin Fraunholz
eng
uncontrolled
acid sphingomyelinase
eng
uncontrolled
staphylococcal alpha-toxin
eng
uncontrolled
sphingomyelinase release
eng
uncontrolled
lysosomal recruitment
eng
uncontrolled
Staphylococcus aureus
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Import
Förderzeitraum 2021
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/24484/fmicb-12-694489.pdf
20411
2020
eng
10
article
1
--
2020-05-12
--
A Role of Sphingosine in the Intracellular Survival of Neisseria gonorrhoeae
Obligate human pathogenic Neisseria gonorrhoeae are the second most frequent bacterial cause of sexually transmitted diseases. These bacteria invade different mucosal tissues and occasionally disseminate into the bloodstream. Invasion into epithelial cells requires the activation of host cell receptors by the formation of ceramide-rich platforms. Here, we investigated the role of sphingosine in the invasion and intracellular survival of gonococci. Sphingosine exhibited an anti-gonococcal activity in vitro. We used specific sphingosine analogs and click chemistry to visualize sphingosine in infected cells. Sphingosine localized to the membrane of intracellular gonococci. Inhibitor studies and the application of a sphingosine derivative indicated that increased sphingosine levels reduced the intracellular survival of gonococci. We demonstrate here, that sphingosine can target intracellular bacteria and may therefore exert a direct bactericidal effect inside cells.
Frontiers in Cellular and Infection Microbiology
2235-2988
10.3389/fcimb.2020.00215
urn:nbn:de:bvb:20-opus-204111
swordwue
2020-05-12T10:39:47+00:00
attachment; filename=deposit.zip
c20a988426404664d7c0c7895457799d
Frontiers in Cellular and Infection Microbiology 2020, 10:215. doi: 10.3389/fcimb.2020.00215
true
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Franziska Solger
Tobias C. Kunz
Julian Fink
Kerstin Paprotka
Pauline Pfister
Franziska Hagen
Fabian Schumacher
Burkhard Kleuser
Jürgen Seibel
Thomas Rudel
eng
uncontrolled
sphingosine
eng
uncontrolled
sphingolipids
eng
uncontrolled
sphingosine kinases
eng
uncontrolled
invasion
eng
uncontrolled
survival
eng
uncontrolled
click chemistry
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Institut für Organische Chemie
Import
Förderzeitraum 2020
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/20411/fcimb-10-00215.pdf
23229
2021
eng
11
article
1
--
--
--
The Expandables: Cracking the Staphylococcal Cell Wall for Expansion Microscopy
Expansion Microscopy (ExM) is a novel tool improving the resolution of fluorescence microscopy by linking the sample into a hydrogel that gets physically expanded in water. Previously, we have used ExM to visualize the intracellular Gram-negative pathogens Chlamydia trachomatis, Simkania negevensis, and Neisseria gonorrhoeae. Gram-positive bacteria have a rigid and thick cell wall that impedes classic expansion strategies. Here we developed an approach, which included a series of enzymatic treatments resulting in isotropic 4× expansion of the Gram-positive pathogen Staphylococcus aureus. We further demonstrate the suitability of the technique for imaging of planktonic bacteria as well as endocytosed, intracellular bacteria at a spatial resolution of approximately 60 nm with conventional confocal laser scanning microscopy.
Frontiers in Cellular and Infection Microbiology
2235-2988
10.3389/fcimb.2021.644750
urn:nbn:de:bvb:20-opus-232292
2021-03-30T06:57:43+00:00
sword
swordwue
attachment; filename=deposit.zip
213b530c49ec8cda1bf76933ec954b68
Frontiers in Cellular and Infection Microbiology 2021, 11:644750. DOI: 10.3389/fcimb.2021.644750
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Tobias C. Kunz
Marcel Rühling
Adriana Moldovan
Kerstin Paprotka
Vera Kozjak-Pavlovic
Thomas Rudel
Martin Fraunholz
eng
uncontrolled
high-resolution imaging
eng
uncontrolled
endosomes
eng
uncontrolled
autophagosomes
eng
uncontrolled
host-pathogen interaction
eng
uncontrolled
expansion microscopy
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Import
Förderzeitraum 2021
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/23229/DataSheet_1.pdf
https://opus.bibliothek.uni-wuerzburg.de/files/23229/fcimb-11-644750-g001.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/fcimb-11-644750-g002.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/fcimb-11-644750-g003.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/fcimb-11-644750-g004.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/Image_1.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/Image_2.tif
https://opus.bibliothek.uni-wuerzburg.de/files/23229/Table_1.docx
https://opus.bibliothek.uni-wuerzburg.de/files/23229/fcimb-11-644750.pdf
20829
2020
eng
8
article
1
--
2020-07-15
--
Using Expansion Microscopy to Visualize and Characterize the Morphology of Mitochondrial Cristae
Mitochondria are double membrane bound organelles indispensable for biological processes such as apoptosis, cell signaling, and the production of many important metabolites, which includes ATP that is generated during the process known as oxidative phosphorylation (OXPHOS). The inner membrane contains folds called cristae, which increase the membrane surface and thus the amount of membrane-bound proteins necessary for the OXPHOS. These folds have been of great interest not only because of their importance for energy conversion, but also because changes in morphology have been linked to a broad range of diseases from cancer, diabetes, neurodegenerative diseases, to aging and infection. With a distance between opposing cristae membranes often below 100 nm, conventional fluorescence imaging cannot provide a resolution sufficient for resolving these structures. For this reason, various highly specialized super-resolution methods including dSTORM, PALM, STED, and SIM have been applied for cristae visualization. Expansion Microscopy (ExM) offers the possibility to perform super-resolution microscopy on conventional confocal microscopes by embedding the sample into a swellable hydrogel that is isotropically expanded by a factor of 4–4.5, improving the resolution to 60–70 nm on conventional confocal microscopes, which can be further increased to ∼ 30 nm laterally using SIM. Here, we demonstrate that the expression of the mitochondrial creatine kinase MtCK linked to marker protein GFP (MtCK-GFP), which localizes to the space between the outer and the inner mitochondrial membrane, can be used as a cristae marker. Applying ExM on mitochondria labeled with this construct enables visualization of morphological changes of cristae and localization studies of mitochondrial proteins relative to cristae without the need for specialized setups. For the first time we present the combination of specific mitochondrial intermembrane space labeling and ExM as a tool for studying internal structure of mitochondria.
Frontiers in Cell and Developmental Biology
2296-634X
10.3389/fcell.2020.00617
urn:nbn:de:bvb:20-opus-208296
swordwue
2020-07-15T05:44:05+00:00
attachment; filename=deposit.zip
979915c25cf5c5c058495285ac32948a
Frontiers in Cell and Developmental Biology 2020, 8:617. doi: 10.3389/fcell.2020.00617
true
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Tobias C. Kunz
Ralph Götz
Shiqiang Gao
Markus Sauer
Vera Kozjak-Pavlovic
eng
uncontrolled
Expansion microscopy
eng
uncontrolled
mitochondria
eng
uncontrolled
cristae
eng
uncontrolled
structured illumination microscope
eng
uncontrolled
ultrastructure
Biowissenschaften; Biologie
open_access
Julius-von-Sachs-Institut für Biowissenschaften
Theodor-Boveri-Institut für Biowissenschaften
Import
Förderzeitraum 2020
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/20829/fcell-08-00617.pdf
20267
2019
eng
e0218303
10
14
article
1
2020-04-09
--
--
Armadillo repeat-containing protein 1 is a dual localization protein associated with mitochondrial intermembrane space bridging complex
Cristae architecture is important for the function of mitochondria, the organelles that play the central role in many cellular processes. The mitochondrial contact site and cristae organizing system (MICOS) together with the sorting and assembly machinery (SAM) forms the mitochondrial intermembrane space bridging complex (MIB), a large protein complex present in mammalian mitochondria that partakes in the formation and maintenance of cristae. We report here a new subunit of the mammalian MICOS/MIB complex, an armadillo repeat-containing protein 1 (ArmC1). ArmC1 localizes both to cytosol and mitochondria, where it associates with the outer mitochondrial membrane through its carboxy-terminus. ArmC1 interacts with other constituents of the MICOS/MIB complex and its amounts are reduced upon MICOS/MIB complex depletion. Mitochondria lacking ArmC1 do not show defects in cristae structure, respiration or protein content, but appear fragmented and with reduced motility. ArmC1 represents therefore a peripheral MICOS/MIB component that appears to play a role in mitochondrial distribution in the cell.
PLoS ONE
10.1371/journal.pone.0218303
urn:nbn:de:bvb:20-opus-202670
PLoS ONE (2019) 14:10, e0218303. https://doi.org/10.1371/journal.pone.0218303
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Fabienne Wagner
Tobias C. Kunz
Suvagata R. Chowdhury
Bernd Thiede
Martin Fraunholz
Debora Eger
Vera Kozjak-Pavlovic
eng
uncontrolled
Mitochondria
eng
uncontrolled
Outer membrane proteins
eng
uncontrolled
HeLa cells
eng
uncontrolled
Immunoprecipitation
eng
uncontrolled
Cytosol
eng
uncontrolled
Small interfering RNAs
eng
uncontrolled
Confocal microscopy
eng
uncontrolled
Cell stainin
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Förderzeitraum 2019
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/20267/Wagner_PLoSONE_2019.pdf
19571
2019
eng
276
9
article
1
--
2019-08-09
--
Detection of chlamydia developmental forms and secreted effectors by expansion microscopy
Expansion microscopy (ExM) is a novel tool to improve the resolution of fluorescence-based microscopy that has not yet been used to visualize intracellular pathogens. Here we show the expansion of the intracellular pathogen Chlamydia trachomatis, enabling to differentiate its two distinct forms, catabolic active reticulate bodies (RB) and infectious elementary bodies (EB), on a conventional confocal microscope. We show that ExM enables the possibility to precisely locate chlamydial effector proteins, such as CPAF or Cdu1, within and outside of the chlamydial inclusion. Thus, we claim that ExM offers the possibility to address a broad range of questions and may be useful for further research on various intracellular pathogens.
Frontiers in Cellular and Infection Microbiology
2235-2988
10.3389/fcimb.2019.00276
urn:nbn:de:bvb:20-opus-195716
Frontiers in Cellular and Infection Microbiology, 2019, 9:276. doi: 10.3389/fcimb.2019.00276
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Tobias C. Kunz
Ralph Götz
Markus Sauer
Thomas Rudel
eng
uncontrolled
expansion microscopy
eng
uncontrolled
chlamydia
eng
uncontrolled
secreted effectors
eng
uncontrolled
developmental forms
eng
uncontrolled
superresolution
eng
uncontrolled
imaging
Biowissenschaften; Biologie
open_access
Theodor-Boveri-Institut für Biowissenschaften
Import
Förderzeitraum 2019
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/19571/fcimb-09-00276.pdf
20175
2019
eng
203
7
article
1
2020-03-16
--
--
Diverse facets of sphingolipid involvement in bacterial infections
Sphingolipids are constituents of the cell membrane that perform various tasks as structural elements and signaling molecules, in addition to regulating many important cellular processes, such as apoptosis and autophagy. In recent years, it has become increasingly clear that sphingolipids and sphingolipid signaling play a vital role in infection processes. In many cases the attachment and uptake of pathogenic bacteria, as well as bacterial development and survival within the host cell depend on sphingolipids. In addition, sphingolipids can serve as antimicrobials, inhibiting bacterial growth and formation of biofilms. This review will give an overview of our current information about these various aspects of sphingolipid involvement in bacterial infections.
Frontiers in Cell and Developmental Biology
10.3389/fcell.2019.00203
urn:nbn:de:bvb:20-opus-201757
Frontiers in Cell and Developmental Biology 2019, 7:203. doi: 10.3389/fcell.2019.00203
false
true
CC BY: Creative-Commons-Lizenz: Namensnennung 4.0 International
Tobias C. Kunz
Vera Kozjak-Pavlovic
eng
uncontrolled
infection
eng
uncontrolled
pathogenic bacteria
eng
uncontrolled
sphingolipids
eng
uncontrolled
ceramide
eng
uncontrolled
autophagy
Medizin und Gesundheit
open_access
Theodor-Boveri-Institut für Biowissenschaften
Förderzeitraum 2019
Universität Würzburg
https://opus.bibliothek.uni-wuerzburg.de/files/20175/Kunz_fcell-07-00203.pdf