Biology subjects

Hof, M.

Publications and source records attributed to Hof, M..

10 recordsLinked to original sources

bioRxiv2025

Modulation of hippocampal synaptic transmission by mast cells at the hippocampal-thalamic border during early brain development

Synaptic development is a highly structured process and can be modulated by the biogenic amine, histamine, whose dysregulation during brain development can be part of the aetiology of the neurodevelopmental disorders Tourettes syndrome and OCD. Well documented in the literature is the extensive histaminergic axonal innervation originating from neurons in tuberomammillary nucleus of the hypothalamus, however, less is known about non-neuronal sources of histamine, such as mast cells, and their potential functional importance in development. Here we set out to investigate the spatiotemporal localisation of mast cells in the developing C57Bl/6 mouse brain and the potential impact of their degranulation on developing synapses. Firstly, we find that the mouse brain contains relatively larger numbers of mast cells during the first and second postnatal week. Secondly, that in this period the highest density of mast cells is found just below the hippocampus at the hippocampal-thalamic border with lower numbers found in other brain regions. Thirdly, using the mast cell degranulator C48/80 we observe that their degranulation can facilitate synaptic transmission at the perforant pathway inputs to the ventral portion of the dentate gyrus which could not be blocked by co-application of the histamine H3 receptor antagonist thioperamide. In contrast histamine superfusion tended to decrease synaptic transmission at this pathway which was sensitive to thioperamide co-application. In conclusion, these results suggest that mast cells are present in the developing C57Bl/6 mouse brain and in particular at the hippocampal-thalamic border where upon degranulation they appear to modulate hippocampal synaptic transmission.

neuroscience↗

bioRxiv2025

Nanoscale PI(4,5)P2 partitioning driven by FGF2 oligomerization triggers membrane pore formation

Fibroblast Growth Factor 2 (FGF2) is a potent tumor cell survival factor. Unlike most secreted proteins, FGF2 lacks a signal peptide and is exported by an unconventional secretory pathway, bypassing the ER/Golgi system. A key event of this unusual secretory mechanism is FGF2 recruitment at the inner plasma membrane leaflet mediated by the phosphoinositide PI(4,5)P2. This interaction induces FGF2 oligomerization which, in turn triggers the formation of a transient lipidic membrane pore with a toroidal structure. FGF2 oligomers populating such membrane pores are captured and disassembled at the outer plasma membrane leaflet through interactions with heparan sulfate chains of the cell surface proteoglycan GPC1, resulting in full translocation of FGF2 across the plasma membrane. Once FGF2 appears on cell surfaces, it engages in signaling complexes initiating autocrine or paracrine signaling. Here, using a multidisciplinary and multiscale approach, we show that FGF2 self-assembles into oligomers that induce lateral PI(4,5)P2 partitioning and membrane remodeling. This process depends on PI(4,5)P2 reaching a critical local threshold concentration that, due to its non-bilayer properties destabilizes the bilayer, facilitating pore formation and FGF2 translocation across the plasma membrane. Our findings highlight a fundamental principle of how protein oligomerization can induce lateral partitioning of membrane lipids that in turn leads to a drastic membrane remodeling event: the transformation of the lipid bilayer into a toroidal membrane pore. The underlying mechanism is likely to be applicable to a wide range of membrane lipid interacting proteins that induce membrane remodeling in other cellular processes.

biochemistry↗

bioRxiv2024

Unraveling the GM1 specificity of Galectin-1 binding to lipid membranes

Galectin-1 (Gal-1) is a galactose-binding protein involved in various cellular functions. Gal-1s activity has been suggested to be connected to two molecular concepts, which are however lacking experimental proof: a) enhanced binding affinity of Gal-1 towards membranes containing monosialotetrahexosylganglioside (GM1) over disialoganglioside GD1a and b) cross-linking of GM1s by homodimers of Gal-1. We provide evidence about the specificity and the nature of Gal-1 interaction with model membranes containing GM1 or GD1a, employing a broad panel of fluorescence-based and label-free experimental techniques, complemented by atomistic biomolecular simulations. Our study demonstrates that Gal-1 binds indeed specifically to GM1, and not to GD1a, when embedded in membranes over a wide range of concentrations (i.e., 30 nM to 10 M). The apparent binding constant is about tens of micromoles. On the other hand, no evidence of Gal-1/GM1 cross-linking was observed. Our findings suggest that cross-linking does not result from sole interactions between GM1 and Gal-1, indicating that in a physiological context, additional triggers are needed, which shift the GM1/Gal-1 equilibria towards the membrane-bound homodimeric Gal-1. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/614102v2_ufig1.gif" ALT="Figure 1"> View larger version (55K): org.highwire.dtl.DTLVardef@c4ff01org.highwire.dtl.DTLVardef@141c82eorg.highwire.dtl.DTLVardef@1bd7c0borg.highwire.dtl.DTLVardef@11af49e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2024

Quantitative imaging of species-specific lipid transport in mammalian cells

Eukaryotic cells produce over 1000 different lipid species which tune organelle membrane properties, control signalling and store energy1,2. How lipid species are selectively sorted between organelles to maintain specific membrane identities is largely unknown due to the difficulty to image lipid transport in cells3. Here, we measured transport and metabolism of individual lipid species in mammalian cells using time-resolved fluorescence imaging of bifunctional lipid probes in combination with ultra-high resolution mass spectrometry and mathematical modelling. Quantification of lipid flux between organelles revealed that directional, non-vesicular lipid transport is responsible for fast, species-selective lipid sorting compared to slow, unspecific vesicular membrane trafficking. Using genetic perturbations, we found that coupling between active lipid flipping and passive non-vesicular transport is a mechanism for directional lipid transport. Comparison of metabolic conversion and transport rates showed that non-vesicular transport dominates the organelle distribution of lipids while species-specific phospholipid metabolism controls neutral lipid accumulation. Our results provide the first quantitative map of retrograde lipid flux in cells4. We anticipate that our pipeline for quantitative mapping of lipid flux through physical and chemical space in cells will boost our understanding of lipids in cell biology and disease.

cell biology↗

bioRxiv2023

Can calmodulin bind to lipids of the cytosolic leaflet of plasma membranes?

Calmodulin (CaM) is a ubiquitous calcium-sensitive messenger in eukaryotic cells. It was previously shown that CaM possesses an affinity for diverse lipid moieties, including those found on CaM-binding proteins. These facts together with our observation that CaM accumulates in membrane-rich protrusions of HeLa cells upon increased cytosolic calcium, motivated us to perform a systematic search for unmediated CaM interactions with model lipid membranes mimicking the cytosolic leaflet of plasma membranes. A range of experimental techniques and Molecular Dynamics simulations proves unambiguously that CaM interacts with lipid bilayers in the presence of calcium ions. Lipids phosphatidylserine (PS) and phosphatidylethanolamine (PE) hold the key to CaM-membrane interactions. Calcium induces an essential conformational rearrangement of CaM, but its binding to the headgroup of PS also neutralizes the membrane negative surface charge. More intriguingly, PE plays a dual role - it forms hydrogen bonds with CaM, but also destabilizes the lipid bilayer to increase exposure of hydrophobic acyl chains to the interacting proteins. Our findings suggest that upon increased intracellular calcium concentration, CaM and the cytosolic leaflet of cellular membranes can be functionally connected.

biophysics↗

bioRxiv2023

Disulfide bridge-dependent dimerization triggers FGF2 membrane translocation into the extracellular space

Fibroblast Growth Factor 2 (FGF2) exits cells by direct translocation across the plasma membrane, a type I pathway of unconventional protein secretion. This process is initiated by PI(4,5)P2-dependent formation of highly dynamic FGF2 oligomers at the inner plasma membrane leaflet, inducing the formation of lipidic membrane pores. Cell surface heparan sulfate chains linked to glypican-1 (GPC1) capture FGF2 at the outer plasma membrane leaflet, completing FGF2 membrane translocation into the extracellular space. While the basic steps of this pathway are well understood, the molecular mechanism by which FGF2 oligomerizes on membrane surfaces remains unclear. In the current study, we demonstrate the initial step of this process to depend on C95-C95 disulfide-bridge-mediated FGF2 dimerization on membrane surfaces, producing the building blocks for higher FGF2 oligomers that drive the formation of membrane pores. We find FGF2 with a C95A substitution to be defective in oligomerization, pore formation, and membrane translocation. Consistently, we demonstrate a C95A variant of FGF2 to be characterized by a severe secretion phenotype. By contrast, while also important for efficient FGF2 secretion from cells, a second cysteine residue on the molecular surface of FGF2 (C77) is not involved in FGF2 oligomerization. Rather, we find C77 to be part of the protein-protein interaction interface through which FGF2 binds to the 1 subunit of the Na,K-ATPase, the landing platform for FGF2 at the inner plasma membrane leaflet. Using cross-linking mass spectrometry, atomistic molecular dynamics simulations combined with a machine learning analysis and cryo-electron tomography, we provide insights into a FGF2 dimerization interface that brings C95 residues in close proximity, resulting in disulfide bridged FGF2 dimers. We propose a mechanism by which they bind with high avidity to PI(4,5)P2 on membrane surfaces. We further propose a tight coupling between FGF2 secretion and the formation of ternary signaling complexes on cell surfaces, hypothesizing that C95-C95 bridged FGF2 dimers are functioning as the molecular units triggering autocrine and paracrine FGF2 signaling.

biochemistry↗

bioRxiv2022

Dissecting the mechanisms of environment sensitivity of smart probes for quantitative assessment of membrane properties

The plasma membrane serves as a crucial platform for a multitude of cellular processes. Its collective biophysical properties are largely determined by the structural diversity of the different lipid species it accommodates. Therefore, a detailed investigation of biophysical properties of the plasma membrane is of utmost importance for a comprehensive understanding of biological processes occurring therein. During the past two decades several environment-sensitive probes have been popular tools to investigate membrane properties. Although these probes are assumed to report on membrane order in similar ways, their individual mechanisms remain to be elucidated. In this study, using model membrane systems, we studied the probes Pro12A, NR12S and NR12A in depth and examined their sensitivity to parameters with potential biological implications, such as the degree of lipid saturation, double bond position and configuration, phospholipid headgroup and cholesterol content. Applying spectral imaging together with atomistic molecular dynamics simulations and time-dependent fluorescent shift analyses, we unravelled individual sensitivities of these probes to different biophysical properties, their distinct localizations and specific relaxation processes in membranes. Overall, Pro12A, NR12S and NR12A serve together as a toolbox with a wide range of applications allowing to select the most appropriate probe for each specific research question.

biophysics↗

bioRxiv2022

No evidence for detectable direct effects of magnetic field on cellular autofluorescence

Dramatically increased levels of electromagnetic radiation in the environment have raised concerns over the potential health hazards of electromagnetic fields. Various biological effects of magnetic fields have been proposed. Despite decades of intensive research, the molecular mechanisms procuring cellular responses remain largely unknown. The current literature is conflicting with regards to evidence that magnetic fields affect functionality directly at cellular level. Therefore, a search for potential direct cellular effects of magnetic fields represents a cornerstone that may propose an explanation for potential health hazards associated with magnetic fields. Recently, it was postulated that autofluorescence of HeLa cells is magnetic field sensitive, relying on single-cell imaging kinetic measurements. Here, we explore the utility of this approach by undertaking a screen for magnetic sensitivity of cellular autofluorescence in statistically relevant numbers (90-107) of HeLa cells. We did not observe any changes in cellular autofluorescence decay, when a modulated magnetic field was applied. We present a number of arguments indicating weak points in the analysis of magnetic field effects based on the imaging of cellular autofluorescence decay. Our work indicates that new methods are required to elucidate the effects of magnetic fields at the cellular level.

cell biology↗

bioRxiv2019

Effect of helical kink in antimicrobial peptides on membrane pore formation

Every cell is protected by a semipermeable membrane. Peptides with the right properties, e.g. Antimicrobial peptides (AMPs), can disrupt this protective barrier by formation of leaky pores. Unfortunately, matching peptide properties with their ability to selectively form pores in bacterial membranes remains elusive. In particular, the proline/glycine kink in helical peptides was reported to both increase and decrease antimicrobial activity. We used computer simulations and fluorescence experiments to show that a kink in helices affects the formation of membrane pores by stabilizing toroidal pores but disrupting barrel-stave pores. The position of the proline/glycine kink in the sequence further controls the specific structure of toroidal pore. Moreover, we demonstrate that two helical peptides can form a kink-like connection with similar behavior as one long helical peptide with a kink. The provided molecular-level insight can be utilized for design and modification of pore forming antibacterial peptides or toxins.

biophysics↗

bioRxiv2016

The impact of a simple α helical transmembrane peptide on lipid membrane dynamics

Transmembrane domains integrate proteins into cellular membranes and support their function. The capacity of these prevalently a-helical structures in mammals to influence membrane properties is poorly understood. Combining experiments with molecular dynamics simulations, we provide evidence that helical transmembrane peptides with their rough surface reduce lateral mobility of membrane constituents. The molecular mechanism involves trapping of lipid acyl chains on the rough surface and segregation of cholesterol from the vicinity of peptides. The observations are supported by our toy model indicating strong effect of rough objects on membrane dynamics. Herein described effect has implications for the organization and function of biological membranes, especially the plasma membrane with high cholesterol content.

biophysics↗