TY - JOUR A1 - Dunsing, Valentin A1 - Petrich, Annett A1 - Chiantia, Salvatore T1 - Multicolor fluorescence fluctuation spectroscopy in living cells via spectral detection JF - eLife N2 - Signaling pathways in biological systems rely on specific interactions between multiple biomolecules. Fluorescence fluctuation spectroscopy provides a powerful toolbox to quantify such interactions directly in living cells. Cross-correlation analysis of spectrally separated fluctuations provides information about intermolecular interactions but is usually limited to two fluorophore species. Here, we present scanning fluorescence spectral correlation spectroscopy (SFSCS), a versatile approach that can be implemented on commercial confocal microscopes, allowing the investigation of interactions between multiple protein species at the plasma membrane. We demonstrate that SFSCS enables cross-talk-free cross-correlation, diffusion, and oligomerization analysis of up to four protein species labeled with strongly overlapping fluorophores. As an example, we investigate the interactions of influenza A virus (IAV) matrix protein 2 with two cellular host factors simultaneously. We furthermore apply raster spectral image correlation spectroscopy for the simultaneous analysis of up to four species and determine the stoichiometry of ternary IAV polymerase complexes in the cell nucleus. KW - fluorescence KW - optical microscopy KW - virus assembly KW - protein-protein KW - interactions KW - diffusion KW - Viruses Y1 - 2021 U6 - https://doi.org/10.7554/eLife.69687 SN - 2050-084X VL - 10 PB - eLife Sciences Publications CY - Cambridge ER - TY - CHAP A1 - Dunsing, Valentin A1 - Petrich, Annett A1 - Chiantia, Salvatore T1 - Spectral detection enables multi-color fluorescence fluctuation spectroscopy studies in living cells BT - Meeting abstract: 65th Annual Meeting of the Biophysical Society (BPS), Feb. 22-26, 2021 T2 - Biophysical journal : BJ / ed. by the Biophysical Society Y1 - 2021 U6 - https://doi.org/10.1016/j.bpj.2020.11.2206 SN - 0006-3495 SN - 1542-0086 VL - 120 IS - 3, Suppl. 1 SP - 356A EP - 356A PB - Cell Press CY - Cambridge ER - TY - JOUR A1 - Petrich, Annett A1 - Dunsing, Valentin A1 - Bobone, Sara A1 - Chiantia, Salvatore T1 - Influenza A M2 recruits M1 to the plasma membrane BT - a fluorescence fluctuation microscopy study JF - Biophysical journal : BJ / ed. by the Biophysical Society N2 - Influenza A virus (IAV) is a respiratory pathogen that causes seasonal epidemics with significant mortality. One of the most abundant proteins in IAV particles is the matrix protein 1 (M1), which is essential for the virus structural stability. M1 organizes virion assembly and budding at the plasma membrane (PM), where it interacts with other viral components. The recruitment of M1 to the PM as well as its interaction with the other viral envelope proteins (hemagglutinin [HA], neuraminidase, matrix protein 2 [M2]) is controversially discussed in previous studies. Therefore, we used fluorescence fluctuation microscopy techniques (i.e., scanning fluorescence cross-correlation spectroscopy and number and brightness) to quantify the oligomeric state of M1 and its interactions with other viral proteins in co-transfected as well as infected cells. Our results indicate that M1 is recruited to the PM by M2, as a consequence of the strong interaction between the two proteins. In contrast, only a weak interaction between M1 and HA was observed. M1-HA interaction occurred only in the event that M1 was already bound to the PM. We therefore conclude that M2 initiates the assembly of IAV by recruiting M1 to the PM, possibly allowing its further interaction with other viral proteins. Y1 - 2021 U6 - https://doi.org/10.1016/j.bpj.2021.11.023 SN - 0006-3495 SN - 1542-0086 VL - 120 IS - 24 SP - 5478 EP - 5490 PB - Cell Press CY - Cambridge ER - TY - THES A1 - Dunsing, Valentin T1 - Fluorescence fluctuation spectroscopy techniques to quantify molecular interactions and dynamics in complex biological systems N2 - Living cells rely on transport and interaction of biomolecules to perform their diverse functions. A powerful toolbox to study these highly dynamic processes in the native environment is provided by fluorescence fluctuation spectroscopy (FFS) techniques. In more detail, FFS takes advantage of the inherent dynamics present in biological systems, such as diffusion, to infer molecular parameters from fluctuations of the signal emitted by an ensemble of fluorescently tagged molecules. In particular, two parameters are accessible: the concentration of molecules and their transit times through the observation volume. In addition, molecular interactions can be measured by analyzing the average signal emitted per molecule - the molecular brightness - and the cross-correlation of signals detected from differently tagged species. In the present work, several FFS techniques were implemented and applied in different biological contexts. In particular, scanning fluorescence correlation spectroscopy (sFCS) was performed to measure protein dynamics and interactions at the plasma membrane (PM) of cells, and number and brightness (N&B) analysis to spatially map molecular aggregation. To account for technical limitations and sample related artifacts, e.g. detector noise, photobleaching, or background signal, several correction schemes were explored. In addition, sFCS was combined with spectral detection and higher moment analysis of the photon count distribution to resolve multiple species at the PM. Using scanning fluorescence cross-correlation spectroscopy and cross-correlation N&B, the interactions of amyloid precursor-like protein 1 (APLP1), a synaptic membrane protein, were investigated. It is shown for the first time directly in living cells, that APLP1 undergoes specific interactions at cell-cell contacts. It is further demonstrated that zinc ions induce formation of large APLP1 clusters that enrich at contact sites and bind to clusters on the opposing cell. Altogether, these results provide direct evidence that APLP1 is a zinc ion dependent neuronal adhesion protein. In the context of APLP1, discrepancies of oligomeric state estimates were observed, which were attributed to non-fluorescent states of the chosen red fluorescent protein (FP) tag mCardinal (mCard). Therefore, multiple FPs and their performance in FFS based measurements of protein interactions were systematically evaluated. The study revealed superior properties of monomeric enhanced green fluorescent protein (mEGFP) and mCherry2. Furthermore, a simple correction scheme allowed unbiased in situ measurements of protein oligomerization by quantifying non-fluorescent state fractions of FP tags. The procedure was experimentally confirmed for biologically relevant protein complexes consisting of up to 12 monomers. In the last part of this work, fluorescence correlation spectroscopy (FCS) and single particle tracking (SPT) were used to characterize diffusive transport dynamics in a bacterial biofilm model. Biofilms are surface adherent bacterial communities, whose structural organization is provided by extracellular polymeric substances (EPS) that form a viscous polymer hydrogel. The presented study revealed a probe size and polymer concentration dependent (anomalous) diffusion hindrance in a reconstituted EPS matrix system caused by polymer chain entanglement at physiological concentrations. This result indicates a meshwork-like organization of the biofilm matrix that allows free diffusion of small particles, but strongly hinders diffusion of larger particles such as bacteriophages. Finally, it is shown that depolymerization of the matrix by phage derived enzymes rapidly facilitated free diffusion. In the context of phage infections, such enzymes may provide a key to evade trapping in the biofilm matrix and promote efficient infection of bacteria. In combination with phage application, matrix depolymerizing enzymes may open up novel antimicrobial strategies against multiresistant bacterial strains, as a promising, more specific alternative to conventional antibiotics. N2 - Die Funktion lebender Zellen basiert auf Transport und Interaktion von Biomolekülen. Zur genauen Untersuchung dieser dynamischen Prozesse in lebenden Zellen eignen sich Fluoreszenzfluktuationsspektroskopieverfahren (FFS). Diese nutzen durch Diffusion oder andere Prozesse auftretende Fluktuationen, um Größen auf molekularer Skala durch statistische Analyse des Signals fluoreszenzmarkierter Moleküle zu ermitteln. Insbesondere können die Konzentration der Moleküle und ihre mittlere Verweildauer im Beobachtungsvolumen quantifiziert werden. Außerdem lassen sich molekulare Interaktionen anhand des mittleren Signals pro Molekül, der sogenannten molekularen Helligkeit, und der Kreuzkorrelation der Signale verschieden markierter Moleküle untersuchen. In der vorliegenden Arbeit wurden verschiedene FFS Methoden etabliert und zur Erforschung biologischer Prozesse genutzt. Um Dynamiken und Bindungsvorgänge an der Zellmembran zu untersuchen, wurde Fluoreszenzkorrelationsspektroskopie (FCS) unter Nutzung eines linearen Scanwegs (sFCS) verwendet. Außerdem wurde die Oligomerisierung von Proteinen mittels Number&Brightness (N&B) Analyse räumlich aufgelöst. Verschiedene Korrekturverfahren wurden validiert und angewandt, um die erhobenen Daten von Störquellen wie Bleichen der Fluorophore oder Hintergrundsignalen zu bereinigen sowie instrumentelle Größen wie Detektionsrauschen zu kalibrieren. Darüber hinaus konnten durch spektral aufgelöste Aufnahme des Fluoreszenzsignals sowie Analyse höherer statistischer Momente mehrere Molekülpopulationen gleichzeitig detektiert werden. Mittels Zweifarben-sFCS und -N&B wurde anschließend das Amyloidvorläuferprotein APLP1 untersucht, welches an Synapsen, den Kontaktstellen von Neuronen, lokalisiert. Mit dem verwendeten Ansatz konnte zum ersten Mal direkt in lebenden Zellen nachgewiesen werden, dass APLP1 spezifische Bindungen an Zellkontaktstellen eingeht. Des Weiteren konnte gezeigt werden, dass Zinkionen eine Anreicherung und verstärkte Interaktion von APLP1 induzieren. Diese Beobachtungen unterstützen die Hypothese, dass APLP1 die Adhäsion benachbarter Zellen vermittelt und diese Funktion konzentrationsabhängig durch Zinkionen reguliert wird. Zur Untersuchung von APLP1 wurde es genetisch mit Fluoreszenzproteinen wie dem rot fluoreszierenden Protein mCardinal fusioniert. Bei der Bestimmung des Oligomerisierungszustands von APLP1 ergaben sich unter Verwendung verschiedener Fluorophore unterschiedliche Ergebnisse. Diese deuteten darauf hin, dass ein Teil der mCardinal Proteine nicht fluoreszierte. Um zu einem tieferen Verständnis dieses Phänomens und dessen Einfluss auf Interaktionsmessungen zu gelangen, wurden häufig verwendete Fluoreszenzproteine systematisch evaluiert. Auf diese Weise konnten zwei Proteine identifiziert werden, grün fluoreszierendes mEGFP und rot fluoreszierendes mCherry2, die den geringsten Anteil an nicht fluoreszierenden Zuständen aufweisen und sich deshalb am besten für Interaktionsmessungen eignen. Mittels eines einfachen Korrekturschemas basierend auf der experimentellen Bestimmung des nicht fluoreszierenden Anteils konnten genaue Messungen des Oligomerisierungszustandes von Proteinen in lebenden Zellen vorgenommen werden, was für biologisch relevante Proteine mit bis zu 12 Untereinheiten erfolgreich gezeigt werden konnte. Im letzten Teil der Arbeit wurden Diffusionsvorgänge in bakteriellen Biofilmen untersucht. Biofilme werden von Bakterienkolonien gebildet, die auf Oberflächen wachsen und beispielsweise zur Verbreitung multiresistenter Keime in Krankenhäusern beitragen. Bei der Bildung von Biofilmen spielen Polymere, die von Bakterien produziert werden, eine entscheidende Rolle. Diese füllen die Zwischenräume im Biofilm mit einer Art Gel, der sogenannten Biofilmmatrix. Anhand von FCS und Einzelpartikelverfolgung konnte gezeigt werden, dass Diffusion von Partikeln in einem rekonstituierten Gel stark von deren Größe sowie der Konzentration der Polymere abhängt. Das untersuchte System bestand hierbei aus langkettigen Zuckermolekülen, die von Biofilmen aufgereinigt wurden und als Modellsystem für die Biofilmmatrix dienten. Im physiologischen Konzentrationsbereich bildete sich ein Polymernetzwerk aus, durch das sich kleine Teilchen frei bewegen konnten, größere Partikel wie z.B. Bakteriophagen jedoch stark verlangsamt wurden. Dies lässt vermuten, dass die Biofilmmatrix die Funktion eines größenabhängigen Filters aufweist. Zersetzung der Polymere mittels Enzymen, die natürlich in Bakteriophagen vorkommen, führte zu freier Diffusion auch größerer Partikel. Die gewonnen Ergebnisse deuten darauf hin, dass solche Enzyme für Phagen eine Schlüsselfunktion besitzen, um Biofilme besser durchdringen und somit Bakterien effizienter infizieren zu können. In Kombination mit Bakteriophagen könnten (zielgerichtet optimierte) Enzyme dieser Art eine vielversprechende, spezifischere Alternative zu konventionellen Antibiotika bei der Bekämpfung multiresistenter Keime darstellen. T2 - Fluoreszenzfluktuationsspektroskopieverfahren zur Bestimmung molekularer Interaktionen und Dynamiken in komplexen biologischen Systemen KW - Fluorescence fluctuation spectroscopy KW - Fluoreszenzfluktuationsspektroskopie KW - Cell-cell adhesion KW - Zell-zell Adhäsion KW - fluorescent proteins KW - Fluoreszenzproteine KW - biofilms KW - Biofilme Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-478494 ER - TY - JOUR A1 - Dunsing, Valentin A1 - Irmscher, Tobias A1 - Barbirz, Stefanie A1 - Chiantia, Salvatore T1 - Purely Polysaccharide-Based Biofilm Matrix Provides Size-Selective Diffusion Barriers for Nanoparticles and Bacteriophages JF - Biomacromolecules : an interdisciplinary journal focused at the interface of polymer science and the biological sciences N2 - Biofilms are complex mixtures of proteins, DNA, and polysaccharides surrounding bacterial communities as protective barriers that can be biochemically modified during the bacterial life cycle. However, their compositional heterogeneity impedes a precise analysis of the contributions of individual matrix components to the biofilm structural organization. To investigate the structural properties of glycan-based biofilms, we analyzed the diffusion dynamics of nanometer-sized objects in matrices of the megadalton-sized anionic polysaccharide, stewartan, the major biofilm component of the plant pathogen, Pantoea stewartii. Fluorescence correlation spectroscopy and single-particle tracking of nanobeads and bacteriophages indicated notable subdiffusive dynamics dependent on probe size and stewartan concentration, in contrast to free diffusion of small molecules. Stewartan enzymatic depolymerization by bacteriophage tailspike proteins rapidly restored unhindered diffusion. We, thus, hypothesize that the glycan polymer stewartan determines the major physicochemical properties of the biofilm, which acts as a selective diffusion barrier for nanometer-sized objects and can be controlled by enzymes. Y1 - 2019 U6 - https://doi.org/10.1021/acs.biomac.9b00938 SN - 1525-7797 SN - 1526-4602 VL - 20 IS - 10 SP - 3842 EP - 3854 PB - American Chemical Society CY - Washington ER - TY - GEN A1 - Dunsing, Valentin A1 - Irmscher, Tobias A1 - Barbirz, Stefanie A1 - Chiantia, Salvatore T1 - Microviscosity of bacterial biofilm matrix characterized by fluorescence correlation spectroscopy and single particle tracking T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2019 U6 - https://doi.org/https://doi.org/10.1007/s00249-019-01373-4 SN - 0175-7571 SN - 1432-1017 VL - 48 SP - S115 EP - S115 PB - Springer CY - New York ER - TY - GEN A1 - Luckner, Madlen A1 - Dunsing, Valentin A1 - Drüke, Markus A1 - Zuehlke, B. A1 - Petazzi, Roberto Arturo A1 - Chiantia, Salvatore A1 - Herrmann, A. T1 - Quantifying protein oligomerization directly in living cells BT - a systematic comparison of fluorescent proteins and application to Influenza A virus infection T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2019 SN - 0175-7571 SN - 1432-1017 VL - 48 SP - S183 EP - S183 PB - Springer CY - New York ER - TY - JOUR A1 - Dunsing, Valentin A1 - Luckner, Madlen A1 - Zuehlke, Boris A1 - Petazzi, Roberto Arturo A1 - Herrmann, Andreas A1 - Chiantia, Salvatore T1 - Optimal fluorescent protein tags for quantifying protein oligomerization in living cells JF - Scientific reports N2 - Fluorescence fluctuation spectroscopy has become a popular toolbox for non-disruptive analysis of molecular interactions in living cells. The quantification of protein oligomerization in the native cellular environment is highly relevant for a detailed understanding of complex biological processes. An important parameter in this context is the molecular brightness, which serves as a direct measure of oligomerization and can be easily extracted from temporal or spatial fluorescence fluctuations. However, fluorescent proteins (FPs) typically used in such studies suffer from complex photophysical transitions and limited maturation, inducing non-fluorescent states. Here, we show how these processes strongly affect molecular brightness measurements. We perform a systematic characterization of non-fluorescent states for commonly used FPs and provide a simple guideline for accurate, unbiased oligomerization measurements in living cells. Further, we focus on novel red FPs and demonstrate that mCherry2, an mCherry variant, possesses superior properties with regards to precise quantification of oligomerization. Y1 - 2018 U6 - https://doi.org/10.1038/s41598-018-28858-0 SN - 2045-2322 VL - 8 PB - Nature Publ. Group CY - London ER - TY - GEN A1 - Luckner, Madlen A1 - Dunsing, Valentin A1 - Chiantia, Salvatore A1 - Hermann, Andreas T1 - Oligomerization and nuclear shuttling dynamics of viral proteins studied by quantitative molecular brightness analysis using fluorescence correlation spectroscopy T2 - Biophysical journal Y1 - 2018 U6 - https://doi.org/10.1016/j.bpj.2017.11.1951 SN - 0006-3495 SN - 1542-0086 VL - 114 IS - 3 SP - 350A EP - 350A PB - Cell Press CY - Cambridge ER - TY - GEN A1 - Dunsing, Valentin A1 - Magnus, Mayer A1 - Liebsch, Filip A1 - Multhaup, Gerhard A1 - Chiantia, Salvatore T1 - Direct Evidence of APLP1 Trans Interactions in Cell-Cell Adhesion Platforms Investigated via Fluorescence Fluctuation Spectroscopy T2 - Biophysical journal N2 - The Amyloid-precursor-like protein 1 (APLP1) is a neuronal type I transmembrane protein which plays a role in synaptic adhesion and synaptogenesis. Past investigations indicated that APLP1 is involved in the formation of protein-protein complexes that bridge the junctions between neighboring cells. Nevertheless, APLP1-APLP1 trans interactions have never been directly observed in higher eukaryotic cells. Here, we investigate APLP1 interactions and dynamics directly in living human embryonic kidney (HEK) cells, using fluorescence fluctuation spectroscopy techniques, namely cross-correlation scanning fluorescence correlation spectroscopy (sFCS) and Number&Brightness (N&B). Our results show that APLP1 forms homotypic trans complexes at cell-cell contacts. In the presence of zinc ions, the protein forms macroscopic clusters, exhibiting an even higher degree of trans binding and strongly reduced dynamics. Further evidence from Giant Plasma Membrane Vesicles and live cell actin staining suggests that the presence of an intact cortical cytoskeleton is required for zinc-induced cis multimerization. Subsequently, large adhesion platforms bridging interacting cells are formed through APLP1-APLP1 direct trans interactions. Taken together, our results provide direct evidence that APLP1 functions as a neuronal zinc-dependent adhesion protein and provide a more detailed understanding of the molecular mechanisms driving the formation of APLP1 adhesion platforms. Further, they show that fluorescence fluctuation spectroscopy techniques are useful tools for the investigation of protein-protein interactions at cell-cell adhesion sites. Y1 - 2018 U6 - https://doi.org/10.1016/j.bpj.2017.11.2067 SN - 0006-3495 SN - 1542-0086 VL - 114 IS - 3 SP - 373A EP - 373A PB - Cell Press CY - Cambridge ER - TY - JOUR A1 - Bobone, Sara A1 - Hilsch, Malte A1 - Storm, Julian A1 - Dunsing, Valentin A1 - Herrmann, Andreas A1 - Chiantia, Salvatore T1 - Phosphatidylserine Lateral Organization Influences the Interaction of Influenza Virus Matrix Protein 1 with Lipid Membranes JF - Journal of virology N2 - Influenza A virus matrix protein 1 (M1) is an essential component involved in the structural stability of the virus and in the budding of new virions from infected cells. A deeper understanding of the molecular basis of virion formation and the budding process is required in order to devise new therapeutic approaches. We performed a detailed investigation of the interaction between M1 and phosphatidylserine (PS) (i.e., its main binding target at the plasma membrane [PM]), as well as the distribution of PS itself, both in model membranes and in living cells. To this end, we used a combination of techniques, including Forster resonance energy transfer (FRET), confocal microscopy imaging, raster image correlation spectroscopy, and number and brightness (N&B) analysis. Our results show that PS can cluster in segregated regions in the plane of the lipid bilayer, both in model bilayers constituted of PS and phosphatidylcholine and in living cells. The viral protein M1 interacts specifically with PS-enriched domains, and such interaction in turn affects its oligomerization process. Furthermore, M1 can stabilize PS domains, as observed in model membranes. For living cells, the presence of PS clusters is suggested by N&B experiments monitoring the clustering of the PS sensor lactadherin. Also, colocalization between M1 and a fluorescent PS probe suggest that, in infected cells, the matrix protein can specifically bind to the regions of PM in which PS is clustered. Taken together, our observations provide novel evidence regarding the role of PS-rich domains in tuning M1-lipid and M1-M1 interactions at the PM of infected cells. IMPORTANCE Influenza virus particles assemble at the plasma membranes (PM) of infected cells. This process is orchestrated by the matrix protein M1, which interacts with membrane lipids while binding to the other proteins and genetic material of the virus. Despite its importance, the initial step in virus assembly (i.e., M1-lipid interaction) is still not well understood. In this work, we show that phosphatidylserine can form lipid domains in physical models of the inner leaflet of the PM. Furthermore, the spatial organization of PS in the plane of the bilayer modulates M1-M1 interactions. Finally, we show that PS domains appear to be present in the PM of living cells and that M1 seems to display a high affinity for them. KW - influenza KW - assembly KW - confocal microscopy KW - fluorescence image analysis KW - lipid rafts KW - matrix protein KW - model membranes KW - phosphatidylserine KW - plasma membrane Y1 - 2017 U6 - https://doi.org/10.1128/JVI.00267-17 SN - 0022-538X SN - 1098-5514 VL - 91 PB - American Society for Microbiology CY - Washington ER - TY - JOUR A1 - Dunsing, Valentin A1 - Mayer, Magnus A1 - Liebsch, Filip A1 - Multhaup, Gerhard A1 - Chiantia, Salvatore T1 - Direct evidence of amyloid precursor-like protein 1 trans interactions in cell-cell adhesion platforms investigated via fluorescence fluctuation spectroscopy JF - Molecular biology of the cell : the official publication of the American Society for Cell Biology N2 - The amyloid precursor-like protein 1 (APLP1) is a type I transmembrane protein that plays a role in synaptic adhesion and synaptogenesis. Past investigations indicated that APLP1 is involved in the formation of protein-protein complexes that bridge the junctions between neighboring cells. Nevertheless, APLP1-APLP1 trans interactions have never been directly observed in higher eukaryotic cells. Here, we investigated APLP1 interactions and dynamics directly in living human embryonic kidney cells using fluorescence fluctuation spectroscopy techniques, namely cross-correlation scanning fluorescence correlation spectroscopy and number and brightness analysis. Our results show that APLP1 forms homotypic trans complexes at cell-cell contacts. In the presence of zinc ions, the protein forms macroscopic clusters, exhibiting an even higher degree of trans binding and strongly reduced dynamics. Further evidence from giant plasma membrane vesicles suggests that the presence of an intact cortical cytoskeleton is required for zinc-induced cis multimerization. Subsequently, large adhesion platforms bridging interacting cells are formed through APLP1-APLP1 trans interactions. Taken together, our results provide direct evidence that APLP1 functions as a neuronal zinc-dependent adhesion protein and allow a more detailed understanding of the molecular mechanisms driving the formation of APLP1 adhesion platforms. Y1 - 2017 U6 - https://doi.org/10.1091/mbc.E17-07-0459 SN - 1059-1524 SN - 1939-4586 VL - 28 SP - 3609 EP - 3620 PB - American Society for Cell Biology CY - Bethesda ER - TY - GEN A1 - Luckner, Madlen A1 - Dunsing, Valentin A1 - Chiantia, Salvatore A1 - Herrmann, Andreas T1 - Influenza virus vRNPs: quantitative investigations via fluorescence cross-correlation spectroscopy T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S368 EP - S368 PB - Springer CY - New York ER - TY - GEN A1 - Dunsing, Valentin A1 - Mayer, M. A1 - Multhaup, G. A1 - Chiantia, Salvatore T1 - Direct visualization of APLP1 cell-cell adhesion platforms via fluorescence fluctuation spectroscopy T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S374 EP - S374 PB - Springer CY - New York ER - TY - JOUR A1 - Mayer, Magnus C. A1 - Schauenburg, Linda A1 - Thompson-Steckel, Greta A1 - Dunsing, Valentin A1 - Kaden, Daniela A1 - Voigt, Philipp A1 - Schaefer, Michael A1 - Chiantia, Salvatore A1 - Kennedy, Timothy E. A1 - Multhaup, Gerhard T1 - Amyloid precursor-like protein 1 (APLP1) exhibits stronger zinc-dependent neuronal adhesion than amyloid precursor protein and APLP2 JF - Journal of neurochemistry N2 - The amyloid precursor protein (APP) and its paralogs, amyloid precursor-like protein 1 (APLP1) and APLP2, are metalloproteins with a putative role both in synaptogenesis and in maintaining synapse structure. Here, we studied the effect of zinc on membrane localization, adhesion, and secretase cleavage of APP, APLP1, and APLP2 in cell culture and rat neurons. For this, we employed live-cell microscopy techniques, a microcontact printing adhesion assay and ELISA for protein detection in cell culture supernatants. We report that zinc induces the multimerization of proteins of the amyloid precursor protein family and enriches them at cellular adhesion sites. Thus, zinc facilitates the formation of de novo APP and APLP1 containing adhesion complexes, whereas it does not have such influence on APLP2. Furthermore, zinc-binding prevented cleavage of APP and APLPs by extracellular secretases. In conclusion, the complexation of zinc modulates neuronal functions of APP and APLPs by (i) regulating formation of adhesion complexes, most prominently for APLP1, and (ii) by reducing the concentrations of neurotrophic soluble APP/APLP ectodomains. KW - amyloid precursor protein KW - amyloid precursor-like protein KW - neuronal adhesion KW - number and brightness KW - zinc Y1 - 2016 U6 - https://doi.org/10.1111/jnc.13540 SN - 0022-3042 SN - 1471-4159 VL - 137 SP - 266 EP - 276 PB - Wiley-Blackwell CY - Hoboken ER -