Biology subjects

Baumgart, T.

Publications and source records attributed to Baumgart, T..

6 recordsLinked to original sources

bioRxiv2026

Multivalency in endocytic accessory proteins orchestrates membrane shape transitions

Fast endophilin-mediated endocytosis (FEME) relies on the coordinated activities of membrane-remodeling proteins and multivalent protein interaction networks. Endophilin is a potent membrane curvature generator through its N-BAR domain and forms multivalent interactions with the proline-rich motifs (PRMs) of lamellipodin via its SH3 domain. Although these interactions are known to promote biomolecular condensation, their functional consequences for membrane remodeling have remained unclear. Here, we investigate how SH3/PRM mediated phase separation regulates endophilin-driven membrane curvature generation. Using purified recombinant proteins, giant unilamellar vesicles, quantitative fluorescence microscopy, and in vitro phase separation assays, we show that multivalent interactions between endophilin and lamellipodin drive the formation of membrane-associated protein-rich phases that strongly suppress endophilin mediated membrane tubulation. Competitive disruption of SH3/PRM interactions with the synaptojanin-derived peptide PP19 (containing only two PRM motifs) rapidly dissolves condensates and restores membrane curvature generation without affecting the intrinsic curvature-generating activity of endophilin. We further demonstrate that this regulatory mechanism is conserved across endophilin A1, A2, and A3 isoforms and depends critically on interaction valency: a multivalent PRM7 construct recapitulates condensate formation and curvature suppression, whereas mono or divalent PRM constructs does not. Together, these findings identify multivalent SH3/PRM interactions as a reversible molecular switch that balances biomolecular condensation and membrane remodeling. Multiple mechanisms, including post-translational modifications, likely regulate the multivalency of these interactions. Our results thus establish a broad functional link between protein phase separation and membrane mechanics, providing a framework for understanding how multivalent interaction networks regulate membrane shaping in multiple endocytic pathways.

biophysics↗

bioRxiv2025

Genetic Code Expansion and Enzymatic Modifications as Accessible Methods for Studying Site-Specific Post-Translational Modifications of Alpha-Synuclein and Tau

Alpha-synuclein (S) and tau play important roles in the pathology of Parkinsons Disease and Alzheimers Disease, respectively, as well as numerous other neurodegenerative diseases. Both proteins are classified as intrinsically disordered proteins (IDPs), as they have no stable structure that underlies their function in healthy tissue, and both proteins are prone to aggregation in disease states. There is substantial interest in understanding the roles that post-translational modifications (PTMs) play in regulating the structural dynamics and function of S and tau monomers, as well as their propensity to aggregate. While there have been many valuable insights into site-specific effects of PTMs garnered through chemical synthesis and semi-synthesis, these techniques are often outside of the expertise of biochemistry and biophysics laboratories wishing to study S and tau. Therefore, we have assembled a primer on genetic code expansion and enzymatic modification approaches to installing PTMs into S and tau site-specifically, including isotopic labeling for NMR and fluorescent labeling for biophysics and microscopy experiments. These methods should be enabling for those wishing to study authentic PTMs in S or tau as well as the broader field of IDPs and aggregating proteins.

biochemistry↗

bioRxiv2025

Strategic Modulation of Polarity and Viscosity Sensitivity of Bimane Molecular Rotor-Based Fluorophores for Imaging α-Synuclein

Molecular rotor-based fluorophores (RBFs) that are target-selective and sensitive to both polarity and viscosity are valuable for diverse biological applications. Here, we have designed next-generation RBFs based on the underexplored bimane fluorophore through either changing in aryl substitution or varying {pi}-linkages between the rotatable electron donors and acceptors to produce red-shifted fluorescence emissions with large Stokes shifts. RBFs exhibit a twisted intramolecular charge transfer mechanism that enables control of polarity and viscosity sensitivity, as well as target selectivity. These features enable their application in: (1) turn-on fluorescent detection of -synuclein (S) fibrils, a hallmark of Parkinsons disease (PD), including amplified fibrils from patient samples; (2) monitoring early misfolding and oligomer formation during S aggregation; and (3) selective imaging of S condensates formed by liquid-liquid phase separation (LLPS). In all three cases, we show that our probes have high levels of selectivity for S versus other aggregating proteins. These properties enable one to study the interplay of S and tau in amyloid aggregation and the mechanisms underlying neurodegenerative disorders.

biochemistry↗

bioRxiv2024

VASP phase separation with priming proteins of fast endophilin mediated endocytosis modulates actin polymerization

Actin polymerization is essential in several clathrin-independent endocytic pathways including fast endophilin mediated endocytosis (FEME), however the actin machinery involved in FEME has been elusive. Here, we show that the actin polymerase VASP colocalizes and interacts directly with the FEME priming complex. We identify Endophilin as a VASP binding partner and establish novel non-canonical interactions between Endophilin and VASP. The major FEME regulators Endophilin and Lamellipodin interact multivalently with VASP to form liquid-like condensates both in solution and on lipid membranes that localize actin polymerization with the extent of actin polymerized regulated by multivalent Endophilin-Lamellipodin interactions. We identify a novel function for Endophilin condensates in bundling actin filaments and show that Endophilin directly binds filamentous actin. Our findings support a model that explains the connection between local actin polymerization and dynamic formation and dissolution of endocytic priming patches in FEME.

biochemistry↗

bioRxiv2021

Multivalent Interactions between Molecular Components Involved in Clathrin Independent Endocytosis Drive Protein Phase Separation

A specific group of transmembrane receptors, including the {beta}1-adrenergic receptor ({beta}1-AR), is internalized through a non-clathrin pathway known as Fast Endophilin Mediated Endocytosis (FEME). A key question is: how does the endocytic machinery assemble and how is it modulated by activated receptors during FEME. Here we show that endophilin, a major regulator of FEME, undergoes a phase transition into liquid-like condensates, which facilitates the formation of multi-protein assemblies by enabling the phase partitioning of endophilin binding proteins. The phase transition can be triggered by specific multivalent binding partners of endophilin in the FEME pathway such as the third intracellular loop (TIL) of the {beta}1-AR, and the C-terminal domain of lamellipodin (LPD). Other endocytic accessory proteins can either partition into, or target interfacial regions of, these condensate droplets. On the membrane, TIL promotes protein clustering in the presence of endophilin and LPD C-terminal domain. Our results demonstrate how the multivalent interactions between endophilin, LPD, and TIL regulate protein assembly formation on the membrane, providing mechanistic insights into the priming and initiation steps of FEME.

biophysics↗

bioRxiv2018

Cell membranes resist flow

The fluid-mosaic model posits a liquid-like plasma membrane, which can flow in response to tension gradients. It is widely assumed that membrane flow transmits local changes in membrane tension across the cell in milliseconds. This conjectured signaling mechanism has been invoked to explain how cells coordinate changes in shape, motility, and vesicle fusion, but the underlying propagation has never been observed. Here we show that propagation of membrane tension occurs quickly in cell-attached blebs, but is largely suppressed in intact cells. The failure of tension to propagate in cells is explained by a fluid dynamical model that incorporates the flow resistance from cytoskeleton-bound transmembrane proteins. In primary endothelial cells, local increases in membrane tension lead only to local activation of mechanosensitive ion channels and to local vesicle fusion. Thus membrane tension is not a mediator of long-range intra-cellular signaling, but local variations in tension mediate distinct processes in sub-cellular domains.

biophysics↗