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 - TY - JOUR A1 - Whiteley, Liam A1 - Haug, Maria A1 - Klein, Kristina A1 - Willmann, Matthias A1 - Bohn, Erwin A1 - Chiantia, Salvatore A1 - Schwarz, Sandra T1 - Cholesterol and host cell surface proteins contribute to cell-cell fusion induced by the Burkholderia type VI secretion system 5 JF - PLoS one N2 - Following escape into the cytoplasm of host cells, Burkholderia pseudomallei and the related species Burkholderia thailandensis employ the type VI secretion system 5 ( T6SS-5) to induce plasma membrane fusion with an adjacent host cell. This process leads to the formation of multinucleated giant cells and facilitates bacterial access to an uninfected host cell in a direct manner. Despite its importance in virulence, the mechanism of the T6SS-5 and the role of host cell factors in cell-cell fusion remain elusive. To date, the T6SS-5 is the only system of bacterial origin known to induce host-cell fusion. To gain insight into the nature of T6SS-5-stimulated membrane fusion, we investigated the contribution of cholesterol and proteins exposed on the host cell surface, which were shown to be critically involved in virus-mediated giant cell formation. In particular, we analyzed the effect of host cell surface protein and cholesterol depletion on the formation of multinucleated giant cells induced by B. thailandensis. Acute protease treatment of RAW264.7 macrophages during infection with B. thailandensis followed by agarose overlay assays revealed a strong reduction in the number of cell-cell fusions compared with EDTA treated cells. Similarly, proteolytic treatment of specifically infected donor cells or uninfected recipient cells significantly decreased multinucleated giant cell formation. Furthermore, modulating host cell cholesterol content by acute cholesterol depletion from cellular membranes by methyl-beta-cyclodextrin treatment or exogenous addition of cholesterol impaired the ability of B. thailandensis to induce cell-cell fusions. The requirement of physiological cholesterol levels suggests that the membrane organization or mechanical properties of the lipid bilayer influence the fusion process. Altogether, our data suggest that membrane fusion induced by B. pseudomallei and B. thailandensis involves a complex interplay between the T6SS-5 and the host cell. Y1 - 2017 U6 - https://doi.org/10.1371/journal.pone.0185715 SN - 1932-6203 VL - 12 PB - PLoS CY - San Fransisco ER - TY - JOUR A1 - Schweighöfer, F. A1 - Moreno, J. A1 - Bobone, Sara A1 - Chiantia, Salvatore A1 - Herrmann, A. A1 - Hecht, S. A1 - Wachtveitl, Josef T1 - Connectivity pattern modifies excited state relaxation dynamics of fluorophore-photoswitch molecular dyads JF - Physical chemistry, chemical physics : a journal of European Chemical Societies N2 - In order to modulate the emission of BODIPY fluorophores, they were connected to a diarylethene (DAE) photoswitch via phenylene-ethynylene linkers of different lengths and orientations. The latter allowed for modulation of the electronic coupling in the prepared four BODIPY-DAE dyads, which were compared also to appropriate BODIPY and DAE model compounds by steady state as well as time-resolved spectroscopies. In their open isomers, all dyads show comparable luminescence behavior indicative of an unperturbed BODIPY fluorophore. In strong contrast, in the closed isomers the BODIPY emission is efficiently quenched but the deactivation mechanism depends on the nature of the linker. The most promising dyad was rendered water-soluble by means of micellar encapsulation and aqueous suspensions were investigated by fluorescence spectroscopy and microscopy. Our results (i) illustrate that the electronic communication between the BODIPY and DAE units can indeed be fine-tuned by the nature of the linker to achieve fluorescence modulation while maintaining photoswitchability and (ii) highlight potential applications to image and control biological processes with high spatio-temporal resolution. Y1 - 2016 U6 - https://doi.org/10.1039/c6cp07112k SN - 1463-9076 SN - 1463-9084 VL - 19 SP - 4010 EP - 4018 PB - Royal Society of Chemistry CY - Cambridge ER - TY - GEN A1 - Höfer, Chris Tina A1 - Di Lella, Santiago A1 - Dahmani, Ismail A1 - Jungnick, Nadine A1 - Bordag, Natalie A1 - Bobone, Sara A1 - Huan, Q. A1 - Keller, S. A1 - Herrmann, A. A1 - Chiantia, Salvatore T1 - Corrigendum to: Structural determinants of the interaction between influenza A virus matrix protein M1 and lipid membranes (Biochimica et Biophysica Acta (BBA) - Biomembranes. - 1861, (2019), pg 1123-1134) T2 - Biochimica et biophysica acta : Biomembranes Y1 - 2019 U6 - https://doi.org/10.1016/j.bbamem.2019.07.002 SN - 0005-2736 SN - 1879-2642 VL - 1861 IS - 10 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Petazzi, Roberto Arturo A1 - Koikkarah Aji, Amit A1 - Tischler, Nicole D. A1 - Chiantia, Salvatore T1 - Detection of envelope glycoprotein assembly from old world hantaviruses in the Golgi apparatus of living cells JF - Journal of virology N2 - Hantaviruses are emerging pathogens that occasionally cause deadly outbreaks in the human population. While the structure of the viral envelope has been characterized with high precision, protein-protein interactions leading to the formation of new virions in infected cells are not fully understood. We used quantitative fluorescence microscopy (i.e., number and brightness analysis and fluorescence fluctuation spectroscopy) to monitor the interactions that lead to oligomeric spike complex formation in the physiological context of living cells. To this aim, we quantified protein-protein interactions for the glycoproteins Gn and Gc from Puumala and Hantaan orthohantaviruses in several cellular models. The oligomerization of each protein was analyzed in relation to subcellular localization, concentration, and the concentration of its interaction partner. Our results indicate that, when expressed separately, Gn and Gc form, respectively, homo-tetrameric and homo-dimeric complexes, in a concentration-dependent manner. Site-directed mutations or deletion mutants showed the specificity of their homotypic interactions. When both glycoproteins were coexpressed, we observed in the Golgi apparatus clear indication of GnGc interactions and the formation of Gn-Gc multimeric protein complexes of different sizes, while using various labeling schemes to minimize the influence of the fluorescent tags. Such large glycoprotein multimers may be identified as multiple Gn viral spikes interconnected via Gc-Gc contacts. This observation provides the possible first evidence for the initial assembly steps of the viral envelope within this organelle, and does so directly in living cells.
IMPORTANCE In this work, we investigate protein-protein interactions that drive the assembly of the hantavirus envelope. These emerging pathogens have the potential to cause deadly outbreaks in the human population. Therefore, it is important to improve our quantitative understanding of the viral assembly process in infected cells, from a molecular point of view. By applying advanced fluorescence microscopy methods, we monitored the formation of viral spike complexes in different cell types. Our data support a model for hantavirus assembly according to which viral spikes are formed via the clustering of hetero-dimers of the two viral glycoproteins Gn and Gc. Furthermore, the observation of large Gn-Gc hetero-multimers provide the possible first evidence for the initial assembly steps of the viral envelope, directly in the Golgi apparatus of living cells. KW - fluorescence fluctuation microscopy KW - number and brightness KW - virus KW - assembly KW - fluorescence correlation spectroscopy KW - protein-protein KW - interaction KW - fluorescence microscopy KW - fluorescent image analysis Y1 - 2021 U6 - https://doi.org/10.1128/JVI.01238-20 SN - 1098-5514 VL - 95 IS - 4 PB - American Society for Microbiology CY - Baltimore, Md. ER - TY - JOUR A1 - Kolyvushko, Oleksandr A1 - Latzke, Juliane A1 - Dahmani, Ismail A1 - Osterrieder, Nikolaus A1 - Chiantia, Salvatore A1 - Azab, Walid T1 - Differentially-charged liposomes interact with alphaherpesviruses and interfere with virus entry JF - Pathogens N2 - Exposure of phosphatidylserine (PS) in the outer leaflet of the plasma membrane is induced by infection with several members of the Alphaherpesvirinae subfamily. There is evidence that PS is used by the equine herpesvirus type 1 (EHV-1) during entry, but the exact role of PS and other phospholipids in the entry process remains unknown. Here, we investigated the interaction of differently charged phospholipids with virus particles and determined their influence on infection. Our data show that liposomes containing negatively charged PS or positively charged DOTAP (N-[1-(2,3-Dioleoyloxy)propyl]-N,N,N-trimethylammonium) inhibited EHV-1 infection, while neutral phosphatidylcholine (PC) had no effect. Inhibition of infection with PS was transient, decreased with time, and was dose dependent. Our findings indicate that both cationic and anionic phospholipids can interact with the virus and reduce infectivity, while, presumably, acting through different mechanisms. Charged phospholipids were found to have antiviral effects and may be used to inhibit EHV-1 infection. KW - alphaherpesvirus KW - EHV-1 KW - phosphatidylserine KW - inhibition KW - pathogen host KW - interaction Y1 - 2020 U6 - https://doi.org/10.3390/pathogens9050359 SN - 2076-0817 VL - 9 IS - 5 PB - MDPI CY - Basel ER - TY - GEN A1 - Kolyvushko, Oleksandr A1 - Latzke, Juliane A1 - Dahmani, Ismail A1 - Osterrieder, Nikolaus A1 - Chiantia, Salvatore A1 - Azab, Walid T1 - Differentially-charged liposomes interact with alphaherpesviruses and interfere with virus entry T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Exposure of phosphatidylserine (PS) in the outer leaflet of the plasma membrane is induced by infection with several members of the Alphaherpesvirinae subfamily. There is evidence that PS is used by the equine herpesvirus type 1 (EHV-1) during entry, but the exact role of PS and other phospholipids in the entry process remains unknown. Here, we investigated the interaction of differently charged phospholipids with virus particles and determined their influence on infection. Our data show that liposomes containing negatively charged PS or positively charged DOTAP (N-[1-(2,3-Dioleoyloxy)propyl]-N,N,N-trimethylammonium) inhibited EHV-1 infection, while neutral phosphatidylcholine (PC) had no effect. Inhibition of infection with PS was transient, decreased with time, and was dose dependent. Our findings indicate that both cationic and anionic phospholipids can interact with the virus and reduce infectivity, while, presumably, acting through different mechanisms. Charged phospholipids were found to have antiviral effects and may be used to inhibit EHV-1 infection. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1088 KW - alphaherpesvirus KW - EHV-1 KW - phosphatidylserine KW - inhibition KW - pathogen host interaction Y1 - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-471895 SN - 1866-8372 IS - 1088 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 - 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 - 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 -