TY - JOUR
AB - Background: This study seeks to evaluate the impact of breast cancer (BRCA) gene status on tumor dissemination pattern, surgical outcome and survival in a multicenter cohort of paired primary ovarian cancer (pOC) and recurrent ovarian cancer (rOC).
Patients and Methods: Medical records and follow-up data from 190 patients were gathered retrospectively. All patients had surgery at pOC and at least one further rOC surgery at four European high-volume centers. Patients were divided into one cohort with confirmed mutation for BRCA1 and/or BRCA2 (BRCAmut) and a second cohort with BRCA wild type or unknown (BRCAwt). Patterns of tumor presentation, surgical outcome and survival data were analyzed between the two groups.
Results: Patients with BRCAmut disease were on average 4 years younger and had significantly more tumor involvement upon diagnosis. Patients with BRCAmut disease showed higher debulking rates at all stages. Multivariate analysis showed that only patient age had significant predictive value for complete tumor resection in pOC. At rOC, however, only BRCAmut status significantly correlated with optimal debulking. Patients with BRCAmut disease showed significantly prolonged overall survival (OS) by 24.3 months. Progression-free survival (PFS) was prolonged in the BRCAmut group at all stages as well, reaching statistical significance during recurrence.
Conclusions: Patients with BRCAmut disease showed a more aggressive course of disease with earlier onset and more extensive tumor dissemination at pOC. However, surgical outcome and OS were significantly better in patients with BRCAmut disease compared with patients with BRCAwt disease. We therefore propose to consider BRCAmut status in regard to patient selection for cytoreductive surgery, especially in rOC.
AU - Glajzer, Jacek
AU - Castillo-Tong, Dan Cacsire
AU - Richter, Rolf
AU - Vergote, Ignace
AU - Kulbe, Hagen
AU - Vanderstichele, Adriaan
AU - Ruscito, Ilary
AU - Trillsch, Fabian
AU - Mustea, Alexander
AU - Kreuzinger, Caroline
AU - Gourley, Charlie
AU - Gabra, Hani
AU - Taube, Eliane T.
AU - Dorigo, Oliver
AU - Horst, David
AU - Keunecke, Carlotta
AU - Baum, Joanna
AU - Angelotti, Timothy
AU - Sehouli, Jalid
AU - Braicu, Elena Ioana
ID - 12205
JF - Annals of Surgical Oncology
KW - Oncology
KW - Surgery
SN - 1068-9265
TI - Impact of BRCA mutation status on tumor dissemination pattern, surgical outcome and patient survival in primary and recurrent high-grade serous ovarian cancer: A multicenter retrospective study by the Ovarian Cancer Therapy-Innovative Models Prolong Survival (OCTIPS) consortium
VL - 30
ER -
TY - JOUR
AU - Glajzer, Jacek
AU - Castillo-Tong, Dan Cacsire
AU - Richter, Rolf
AU - Vergote, Ignace
AU - Kulbe, Hagen
AU - Vanderstichele, Adriaan
AU - Ruscito, Ilary
AU - Trillsch, Fabian
AU - Mustea, Alexander
AU - Kreuzinger, Caroline
AU - Gourley, Charlie
AU - Gabra, Hani
AU - Taube, Eliane T.
AU - Dorigo, Oliver
AU - Horst, David
AU - Keunecke, Carlotta
AU - Baum, Joanna
AU - Angelotti, Timothy
AU - Sehouli, Jalid
AU - Braicu, Elena Ioana
ID - 12115
JF - Annals of Surgical Oncology
KW - Oncology
KW - Surgery
SN - 1068-9265
TI - ASO Visual Abstract: Impact of BRCA mutation status on tumor dissemination pattern, surgical outcome, and patient survival in primary and recurrent high-grade serous ovarian cancer (HGSOC). A multicenter, retrospective study of the ovarian cancer therapy—innovative models prolong survival (OCTIPS) consortium
VL - 30
ER -
TY - DATA
AB - 3D-reconstruction of living brain tissue down to individual synapse level would create opportunities for decoding the dynamics and structure-function relationships of the brain’s complex and dense information processing network. However, it has been hindered by insufficient 3D-resolution, inadequate signal-to-noise-ratio, and prohibitive light burden in optical imaging, whereas electron microscopy is inherently static. Here we solved these challenges by developing an integrated optical/machine learning technology, LIONESS (Live Information-Optimized Nanoscopy Enabling Saturated Segmentation). It leverages optical modifications to stimulated emission depletion (STED) microscopy in comprehensively, extracellularly labelled tissue and prior information on sample structure via machine learning to simultaneously achieve isotropic super-resolution, high signal-to-noise-ratio, and compatibility with living tissue. This allows dense deep-learning-based instance segmentation and 3D-reconstruction at synapse level incorporating molecular, activity, and morphodynamic information. LIONESS opens up avenues for studying the dynamic functional (nano-)architecture of living brain tissue.
AU - Danzl, Johann G
ID - 12817
TI - Research data for the publication "Dense 4D nanoscale reconstruction of living brain tissue"
ER -
TY - JOUR
AB - Three-dimensional (3D) reconstruction of living brain tissue down to an individual synapse level would create opportunities for decoding the dynamics and structure–function relationships of the brain’s complex and dense information processing network; however, this has been hindered by insufficient 3D resolution, inadequate signal-to-noise ratio and prohibitive light burden in optical imaging, whereas electron microscopy is inherently static. Here we solved these challenges by developing an integrated optical/machine-learning technology, LIONESS (live information-optimized nanoscopy enabling saturated segmentation). This leverages optical modifications to stimulated emission depletion microscopy in comprehensively, extracellularly labeled tissue and previous information on sample structure via machine learning to simultaneously achieve isotropic super-resolution, high signal-to-noise ratio and compatibility with living tissue. This allows dense deep-learning-based instance segmentation and 3D reconstruction at a synapse level, incorporating molecular, activity and morphodynamic information. LIONESS opens up avenues for studying the dynamic functional (nano-)architecture of living brain tissue.
AU - Velicky, Philipp
AU - Miguel Villalba, Eder
AU - Michalska, Julia M
AU - Lyudchik, Julia
AU - Wei, Donglai
AU - Lin, Zudi
AU - Watson, Jake
AU - Troidl, Jakob
AU - Beyer, Johanna
AU - Ben Simon, Yoav
AU - Sommer, Christoph M
AU - Jahr, Wiebke
AU - Cenameri, Alban
AU - Broichhagen, Johannes
AU - Grant, Seth G.N.
AU - Jonas, Peter M
AU - Novarino, Gaia
AU - Pfister, Hanspeter
AU - Bickel, Bernd
AU - Danzl, Johann G
ID - 13267
JF - Nature Methods
SN - 1548-7091
TI - Dense 4D nanoscale reconstruction of living brain tissue
VL - 20
ER -
TY - JOUR
AB - We developed LIONESS, a technology that leverages improvements to optical super-resolution microscopy and prior information on sample structure via machine learning to overcome the limitations (in 3D-resolution, signal-to-noise ratio and light exposure) of optical microscopy of living biological specimens. LIONESS enables dense reconstruction of living brain tissue and morphodynamics visualization at the nanoscale.
AU - Danzl, Johann G
AU - Velicky, Philipp
ID - 14770
IS - 8
JF - Nature Methods
KW - Cell Biology
KW - Molecular Biology
KW - Biochemistry
KW - Biotechnology
SN - 1548-7091
TI - LIONESS enables 4D nanoscale reconstruction of living brain tissue
VL - 20
ER -
TY - DATA
AB - Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here, we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanometer synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS uses fixation-compatible extracellular labeling and optical imaging, including stimulated emission depletion or expansion microscopy, to comprehensively delineate cellular structures. It enables three-dimensional reconstruction of single synapses and mapping of synaptic connectivity by identification and analysis of putative synaptic cleft regions. Applying CATS to the mouse hippocampal mossy fiber circuitry, we reconstructed and quantified the synaptic input and output structure of identified neurons. We furthermore demonstrate applicability to clinically derived human tissue samples, including formalin-fixed paraffin-embedded routine diagnostic specimens, for visualizing the cellular architecture of brain tissue in health and disease.
AU - Danzl, Johann G
ID - 13126
TI - Research data for the publication "Imaging brain tissue architecture across millimeter to nanometer scales"
ER -
TY - DATA
AB - The emergence of large-scale order in self-organized systems relies on local interactions between individual components. During bacterial cell division, FtsZ -- a prokaryotic homologue of the eukaryotic protein tubulin -- polymerizes into treadmilling filaments that further organize into a cytoskeletal ring. In vitro, FtsZ filaments can form dynamic chiral assemblies. However, how the active and passive properties of individual filaments relate to these large-scale self-organized structures remains poorly understood. Here, we connect single filament properties with the mesoscopic scale by combining minimal active matter simulations and biochemical reconstitution experiments. We show that density and flexibility of active chiral filaments define their global order. At intermediate densities, curved, flexible filaments organize into chiral rings and polar bands. An effectively nematic organization dominates for high densities and for straight, mutant filaments with increased rigidity. Our predicted phase diagram captures these features quantitatively, demonstrating how the flexibility, density and chirality of active filaments affect their collective behaviour. Our findings shed light on the fundamental properties of active chiral matter and explain how treadmilling FtsZ filaments organize during bacterial cell division.
AU - Dunajova, Zuzana
AU - Prats Mateu, Batirtze
AU - Radler, Philipp
AU - Lim, Keesiang
AU - Brandis, Dörte
AU - Velicky, Philipp
AU - Danzl, Johann G
AU - Wong, Richard W.
AU - Elgeti, Jens
AU - Hannezo, Edouard B
AU - Loose, Martin
ID - 13116
TI - Chiral and nematic phases of flexible active filaments
ER -
TY - JOUR
AB - The emergence of large-scale order in self-organized systems relies on local interactions between individual components. During bacterial cell division, FtsZ—a prokaryotic homologue of the eukaryotic protein tubulin—polymerizes into treadmilling filaments that further organize into a cytoskeletal ring. In vitro, FtsZ filaments can form dynamic chiral assemblies. However, how the active and passive properties of individual filaments relate to these large-scale self-organized structures remains poorly understood. Here we connect single-filament properties with the mesoscopic scale by combining minimal active matter simulations and biochemical reconstitution experiments. We show that the density and flexibility of active chiral filaments define their global order. At intermediate densities, curved, flexible filaments organize into chiral rings and polar bands. An effectively nematic organization dominates for high densities and for straight, mutant filaments with increased rigidity. Our predicted phase diagram quantitatively captures these features, demonstrating how the flexibility, density and chirality of the active filaments affect their collective behaviour. Our findings shed light on the fundamental properties of active chiral matter and explain how treadmilling FtsZ filaments organize during bacterial cell division.
AU - Dunajova, Zuzana
AU - Prats Mateu, Batirtze
AU - Radler, Philipp
AU - Lim, Keesiang
AU - Brandis, Dörte
AU - Velicky, Philipp
AU - Danzl, Johann G
AU - Wong, Richard W.
AU - Elgeti, Jens
AU - Hannezo, Edouard B
AU - Loose, Martin
ID - 13314
JF - Nature Physics
SN - 1745-2473
TI - Chiral and nematic phases of flexible active filaments
VL - 19
ER -
TY - JOUR
AB - Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanometer synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS uses fixation-compatible extracellular labeling and optical imaging, including stimulated emission depletion or expansion microscopy, to comprehensively delineate cellular structures. It enables three-dimensional reconstruction of single synapses and mapping of synaptic connectivity by identification and analysis of putative synaptic cleft regions. Applying CATS to the mouse hippocampal mossy fiber circuitry, we reconstructed and quantified the synaptic input and output structure of identified neurons. We furthermore demonstrate applicability to clinically derived human tissue samples, including formalin-fixed paraffin-embedded routine diagnostic specimens, for visualizing the cellular architecture of brain tissue in health and disease.
AU - Michalska, Julia M
AU - Lyudchik, Julia
AU - Velicky, Philipp
AU - Korinkova, Hana
AU - Watson, Jake
AU - Cenameri, Alban
AU - Sommer, Christoph M
AU - Amberg, Nicole
AU - Venturino, Alessandro
AU - Roessler, Karl
AU - Czech, Thomas
AU - Höftberger, Romana
AU - Siegert, Sandra
AU - Novarino, Gaia
AU - Jonas, Peter M
AU - Danzl, Johann G
ID - 14257
JF - Nature Biotechnology
SN - 1087-0156
TI - Imaging brain tissue architecture across millimeter to nanometer scales
ER -
TY - THES
AB - The brain is an exceptionally sophisticated organ consisting of billions of cells and trillions of
connections that orchestrate our cognition and behavior. To decode its complex connectivity, it is
pivotal to disentangle its intricate architecture spanning from cm-sized circuits down to tens of
nm-small synapses.
To achieve this goal, I developed CATS – Comprehensive Analysis of nervous Tissue across
Scales, a versatile toolbox for obtaining a holistic view of nervous tissue context with (superresolution) fluorescence microscopy. CATS combines comprehensive labeling of the extracellular
space, that is compatible with chemical fixation, with information on molecular markers, superresolved data acquisition and machine-learning based data analysis for segmentation and synapse
identification.
I used CATS to analyze key features of nervous tissue connectivity, ranging from whole tissue
architecture, neuronal in- and output-fields, down to synapse morphology.
Focusing on the hippocampal circuitry, I quantified synaptic transmission properties of mossy
fiber boutons and analyzed the connectivity pattern of dentate gyrus granule cells with CA3
pyramidal neurons. This shows that CATS is a viable tool to study hallmarks of neuronal
connectivity with light microscopy.
AU - Michalska, Julia M
ID - 12470
SN - 2663-337X
TI - A versatile toolbox for the comprehensive analysis of nervous tissue organization with light microscopy
ER -
TY - JOUR
AB - The mammalian hippocampal formation (HF) plays a key role in several higher brain functions, such as spatial coding, learning and memory. Its simple circuit architecture is often viewed as a trisynaptic loop, processing input originating from the superficial layers of the entorhinal cortex (EC) and sending it back to its deeper layers. Here, we show that excitatory neurons in layer 6b of the mouse EC project to all sub-regions comprising the HF and receive input from the CA1, thalamus and claustrum. Furthermore, their output is characterized by unique slow-decaying excitatory postsynaptic currents capable of driving plateau-like potentials in their postsynaptic targets. Optogenetic inhibition of the EC-6b pathway affects spatial coding in CA1 pyramidal neurons, while cell ablation impairs not only acquisition of new spatial memories, but also degradation of previously acquired ones. Our results provide evidence of a functional role for cortical layer 6b neurons in the adult brain.
AU - Ben Simon, Yoav
AU - Käfer, Karola
AU - Velicky, Philipp
AU - Csicsvari, Jozsef L
AU - Danzl, Johann G
AU - Jonas, Peter M
ID - 11951
JF - Nature Communications
KW - General Physics and Astronomy
KW - General Biochemistry
KW - Genetics and Molecular Biology
KW - General Chemistry
KW - Multidisciplinary
SN - 2041-1723
TI - A direct excitatory projection from entorhinal layer 6b neurons to the hippocampus contributes to spatial coding and memory
VL - 13
ER -
TY - GEN
AB - Complex wiring between neurons underlies the information-processing network enabling all brain functions, including cognition and memory. For understanding how the network is structured, processes information, and changes over time, comprehensive visualization of the architecture of living brain tissue with its cellular and molecular components would open up major opportunities. However, electron microscopy (EM) provides nanometre-scale resolution required for full in-silico reconstruction1–5, yet is limited to fixed specimens and static representations. Light microscopy allows live observation, with super-resolution approaches6–12 facilitating nanoscale visualization, but comprehensive 3D-reconstruction of living brain tissue has been hindered by tissue photo-burden, photobleaching, insufficient 3D-resolution, and inadequate signal-to-noise ratio (SNR). Here we demonstrate saturated reconstruction of living brain tissue. We developed an integrated imaging and analysis technology, adapting stimulated emission depletion (STED) microscopy6,13 in extracellularly labelled tissue14 for high SNR and near-isotropic resolution. Centrally, a two-stage deep-learning approach leveraged previously obtained information on sample structure to drastically reduce photo-burden and enable automated volumetric reconstruction down to single synapse level. Live reconstruction provides unbiased analysis of tissue architecture across time in relation to functional activity and targeted activation, and contextual understanding of molecular labelling. This adoptable technology will facilitate novel insights into the dynamic functional architecture of living brain tissue.
AU - Velicky, Philipp
AU - Miguel Villalba, Eder
AU - Michalska, Julia M
AU - Wei, Donglai
AU - Lin, Zudi
AU - Watson, Jake
AU - Troidl, Jakob
AU - Beyer, Johanna
AU - Ben Simon, Yoav
AU - Sommer, Christoph M
AU - Jahr, Wiebke
AU - Cenameri, Alban
AU - Broichhagen, Johannes
AU - Grant, Seth G. N.
AU - Jonas, Peter M
AU - Novarino, Gaia
AU - Pfister, Hanspeter
AU - Bickel, Bernd
AU - Danzl, Johann G
ID - 11943
T2 - bioRxiv
TI - Saturated reconstruction of living brain tissue
ER -
TY - GEN
AB - Mapping the complex and dense arrangement of cells and their connectivity in brain tissue demands nanoscale spatial resolution imaging. Super-resolution optical microscopy excels at visualizing specific molecules and individual cells but fails to provide tissue context. Here we developed Comprehensive Analysis of Tissues across Scales (CATS), a technology to densely map brain tissue architecture from millimeter regional to nanoscopic synaptic scales in diverse chemically fixed brain preparations, including rodent and human. CATS leverages fixation-compatible extracellular labeling and advanced optical readout, in particular stimulated-emission depletion and expansion microscopy, to comprehensively delineate cellular structures. It enables 3D-reconstructing single synapses and mapping synaptic connectivity by identification and tailored analysis of putative synaptic cleft regions. Applying CATS to the hippocampal mossy fiber circuitry, we demonstrate its power to reveal the system’s molecularly informed ultrastructure across spatial scales and assess local connectivity by reconstructing and quantifying the synaptic input and output structure of identified neurons.
AU - Michalska, Julia M
AU - Lyudchik, Julia
AU - Velicky, Philipp
AU - Korinkova, Hana
AU - Watson, Jake
AU - Cenameri, Alban
AU - Sommer, Christoph M
AU - Venturino, Alessandro
AU - Roessler, Karl
AU - Czech, Thomas
AU - Siegert, Sandra
AU - Novarino, Gaia
AU - Jonas, Peter M
AU - Danzl, Johann G
ID - 11950
T2 - bioRxiv
TI - Uncovering brain tissue architecture across scales with super-resolution light microscopy
ER -
TY - JOUR
AB - Mutations in the chromodomain helicase DNA-binding 8 (CHD8) gene are a frequent cause of autism spectrum disorder (ASD). While its phenotypic spectrum often encompasses macrocephaly, implicating cortical abnormalities, how CHD8 haploinsufficiency affects neurodevelopmental is unclear. Here, employing human cerebral organoids, we find that CHD8 haploinsufficiency disrupted neurodevelopmental trajectories with an accelerated and delayed generation of, respectively, inhibitory and excitatory neurons that yields, at days 60 and 120, symmetrically opposite expansions in their proportions. This imbalance is consistent with an enlargement of cerebral organoids as an in vitro correlate of patients’ macrocephaly. Through an isogenic design of patient-specific mutations and mosaic organoids, we define genotype-phenotype relationships and uncover their cell-autonomous nature. Our results define cell-type-specific CHD8-dependent molecular defects related to an abnormal program of proliferation and alternative splicing. By identifying cell-type-specific effects of CHD8 mutations, our study uncovers reproducible developmental alterations that may be employed for neurodevelopmental disease modeling.
AU - Villa, Carlo Emanuele
AU - Cheroni, Cristina
AU - Dotter, Christoph
AU - López-Tóbon, Alejandro
AU - Oliveira, Bárbara
AU - Sacco, Roberto
AU - Yahya, Aysan Çerağ
AU - Morandell, Jasmin
AU - Gabriele, Michele
AU - Tavakoli, Mojtaba
AU - Lyudchik, Julia
AU - Sommer, Christoph M
AU - Gabitto, Mariano
AU - Danzl, Johann G
AU - Testa, Giuseppe
AU - Novarino, Gaia
ID - 11160
IS - 1
JF - Cell Reports
KW - General Biochemistry
KW - Genetics and Molecular Biology
SN - 2211-1247
TI - CHD8 haploinsufficiency links autism to transient alterations in excitatory and inhibitory trajectories
VL - 39
ER -
TY - CHAP
AB - Expansion microscopy is a recently developed super-resolution imaging technique, which provides an alternative to optics-based methods such as deterministic approaches (e.g. STED) or stochastic approaches (e.g. PALM/STORM). The idea behind expansion microscopy is to embed the biological sample in a swellable gel, and then to expand it isotropically, thereby increasing the distance between the fluorophores. This approach breaks the diffraction barrier by simply separating the emission point-spread-functions of the fluorophores. The resolution attainable in expansion microscopy is thus directly dependent on the separation that can be achieved, i.e. on the expansion factor. The original implementation of the technique achieved an expansion factor of fourfold, for a resolution of 70–80 nm. The subsequently developed X10 method achieves an expansion factor of 10-fold, for a resolution of 25–30 nm. This technique can be implemented with minimal technical requirements on any standard fluorescence microscope, and is more easily applied for multi-color imaging than either deterministic or stochastic super-resolution approaches. This renders X10 expansion microscopy a highly promising tool for new biological discoveries, as discussed here, and as demonstrated by several recent applications.
AU - Truckenbrodt, Sven M
AU - Rizzoli, Silvio O.
ID - 7941
SN - 0091-679X
T2 - Methods in Cell Biology
TI - Simple multi-color super-resolution by X10 microscopy
VL - 161
ER -
TY - JOUR
AU - Pinkard, Henry
AU - Stuurman, Nico
AU - Ivanov, Ivan E.
AU - Anthony, Nicholas M.
AU - Ouyang, Wei
AU - Li, Bin
AU - Yang, Bin
AU - Tsuchida, Mark A.
AU - Chhun, Bryant
AU - Zhang, Grace
AU - Mei, Ryan
AU - Anderson, Michael
AU - Shepherd, Douglas P.
AU - Hunt-Isaak, Ian
AU - Dunn, Raymond L.
AU - Jahr, Wiebke
AU - Kato, Saul
AU - Royer, Loïc A.
AU - Thiagarajah, Jay R.
AU - Eliceiri, Kevin W.
AU - Lundberg, Emma
AU - Mehta, Shalin B.
AU - Waller, Laura
ID - 9258
IS - 3
JF - Nature Methods
SN - 1548-7091
TI - Pycro-Manager: Open-source software for customized and reproducible microscope control
VL - 18
ER -
TY - JOUR
AB - De novo loss of function mutations in the ubiquitin ligase-encoding gene Cullin3 lead to autism spectrum disorder (ASD). In mouse, constitutive haploinsufficiency leads to motor coordination deficits as well as ASD-relevant social and cognitive impairments. However, induction of Cul3 haploinsufficiency later in life does not lead to ASD-relevant behaviors, pointing to an important role of Cul3 during a critical developmental window. Here we show that Cul3 is essential to regulate neuronal migration and, therefore, constitutive Cul3 heterozygous mutant mice display cortical lamination abnormalities. At the molecular level, we found that Cul3 controls neuronal migration by tightly regulating the amount of Plastin3 (Pls3), a previously unrecognized player of neural migration. Furthermore, we found that Pls3 cell-autonomously regulates cell migration by regulating actin cytoskeleton organization, and its levels are inversely proportional to neural migration speed. Finally, we provide evidence that cellular phenotypes associated with autism-linked gene haploinsufficiency can be rescued by transcriptional activation of the intact allele in vitro, offering a proof of concept for a potential therapeutic approach for ASDs.
AU - Morandell, Jasmin
AU - Schwarz, Lena A
AU - Basilico, Bernadette
AU - Tasciyan, Saren
AU - Dimchev, Georgi A
AU - Nicolas, Armel
AU - Sommer, Christoph M
AU - Kreuzinger, Caroline
AU - Dotter, Christoph
AU - Knaus, Lisa
AU - Dobler, Zoe
AU - Cacci, Emanuele
AU - Schur, Florian KM
AU - Danzl, Johann G
AU - Novarino, Gaia
ID - 9429
IS - 1
JF - Nature Communications
KW - General Biochemistry
KW - Genetics and Molecular Biology
TI - Cul3 regulates cytoskeleton protein homeostasis and cell migration during a critical window of brain development
VL - 12
ER -
TY - JOUR
AB - Super-resolution fluorescence microscopy has become an important catalyst for discovery in the life sciences. In STimulated Emission Depletion (STED) microscopy, a pattern of light drives fluorophores from a signal-emitting on-state to a non-signalling off-state. Only emitters residing in a sub-diffraction volume around an intensity minimum are allowed to fluoresce, rendering them distinguishable from the nearby, but dark fluorophores. STED routinely achieves resolution in the few tens of nanometers range in biological samples and is suitable for live imaging. Here, we review the working principle of STED and provide general guidelines for successful STED imaging. The strive for ever higher resolution comes at the cost of increased light burden. We discuss techniques to reduce light exposure and mitigate its detrimental effects on the specimen. These include specialized illumination strategies as well as protecting fluorophores from photobleaching mediated by high-intensity STED light. This opens up the prospect of volumetric imaging in living cells and tissues with diffraction-unlimited resolution in all three spatial dimensions.
AU - Jahr, Wiebke
AU - Velicky, Philipp
AU - Danzl, Johann G
ID - 6808
IS - 3
JF - Methods
SN - 1046-2023
TI - Strategies to maximize performance in STimulated Emission Depletion (STED) nanoscopy of biological specimens
VL - 174
ER -
TY - JOUR
AB - Acute brain slice preparation is a powerful experimental model for investigating the characteristics of synaptic function in the brain. Although brain tissue is usually cut at ice-cold temperature (CT) to facilitate slicing and avoid neuronal damage, exposure to CT causes molecular and architectural changes of synapses. To address these issues, we investigated ultrastructural and electrophysiological features of synapses in mouse acute cerebellar slices prepared at ice-cold and physiological temperature (PT). In the slices prepared at CT, we found significant spine loss and reconstruction, synaptic vesicle rearrangement and decrease in synaptic proteins, all of which were not detected in slices prepared at PT. Consistent with these structural findings, slices prepared at PT showed higher release probability. Furthermore, preparation at PT allows electrophysiological recording immediately after slicing resulting in higher detectability of long-term depression (LTD) after motor learning compared with that at CT. These results indicate substantial advantages of the slice preparation at PT for investigating synaptic functions in different physiological conditions.
AU - Eguchi, Kohgaku
AU - Velicky, Philipp
AU - Hollergschwandtner, Elena
AU - Itakura, Makoto
AU - Fukazawa, Yugo
AU - Danzl, Johann G
AU - Shigemoto, Ryuichi
ID - 7665
JF - Frontiers in Cellular Neuroscience
SN - 16625102
TI - Advantages of acute brain slices prepared at physiological temperature in the characterization of synaptic functions
VL - 14
ER -
TY - GEN
AB - De novo loss of function mutations in the ubiquitin ligase-encoding gene Cullin3 (CUL3) lead to autism spectrum disorder (ASD). Here, we used Cul3 mouse models to evaluate the consequences of Cul3 mutations in vivo. Our results show that Cul3 haploinsufficient mice exhibit deficits in motor coordination as well as ASD-relevant social and cognitive impairments. Cul3 mutant brain displays cortical lamination abnormalities due to defective neuronal migration and reduced numbers of excitatory and inhibitory neurons. In line with the observed abnormal columnar organization, Cul3 haploinsufficiency is associated with decreased spontaneous excitatory and inhibitory activity in the cortex. At the molecular level, employing a quantitative proteomic approach, we show that Cul3 regulates cytoskeletal and adhesion protein abundance in mouse embryos. Abnormal regulation of cytoskeletal proteins in Cul3 mutant neuronal cells results in atypical organization of the actin mesh at the cell leading edge, likely causing the observed migration deficits. In contrast to these important functions early in development, Cul3 deficiency appears less relevant at adult stages. In fact, induction of Cul3 haploinsufficiency in adult mice does not result in the behavioral defects observed in constitutive Cul3 haploinsufficient animals. Taken together, our data indicate that Cul3 has a critical role in the regulation of cytoskeletal proteins and neuronal migration and that ASD-associated defects and behavioral abnormalities are primarily due to Cul3 functions at early developmental stages.
AU - Morandell, Jasmin
AU - Schwarz, Lena A
AU - Basilico, Bernadette
AU - Tasciyan, Saren
AU - Nicolas, Armel
AU - Sommer, Christoph M
AU - Kreuzinger, Caroline
AU - Knaus, Lisa
AU - Dobler, Zoe
AU - Cacci, Emanuele
AU - Danzl, Johann G
AU - Novarino, Gaia
ID - 7800
T2 - bioRxiv
TI - Cul3 regulates cytoskeleton protein homeostasis and cell migration during a critical window of brain development
ER -