Biology subjects

Hendricks, A. G.

Publications and source records attributed to Hendricks, A. G..

17 recordsLinked to original sources

bioRxiv2026

SARS-CoV-2 maturation driven by a mechano-active nucleocapsid-RNA condensate

The nucleocapsid (N) protein, the core component of the SARS-CoV-2 virus, binds to the 30-kb viral genomic RNA (vgRNA) to form ribonucleoprotein assemblies that are packaged into ~100 nm membrane-enclosed virions. In addition to viral assembly, the N protein performs other functions, including roles in viral mRNA transcription, replication, and immune regulation, making it a key target for developing diagnostics and vaccines. Recent studies show that N protein and RNA undergo phase separation to form a biomolecular condensate that constitutes the viral core. However, how this condensate becomes selectively enveloped by a membrane remains unclear. Here, using a minimal reconstituted system, we demonstrate that N protein-vgRNA condensates are spontaneously enveloped by lipid bilayer membranes, whereas condensates formed by N protein alone or with genomic RNA fragments fail to undergo envelopment and instead adhere to or weakly deform membranes. We show that RNA length tunes the material properties of N protein condensates, with vgRNA imparting enhanced elasticity. We propose a physical model for SARS-CoV-2 maturation in which adhesion between the membrane and the N protein-vgRNA condensate drives membrane bending, while condensate elasticity preserves core geometry. These results identify RNA-dependent material properties as an underlying physical principle for selective SARS-CoV-2 maturation.

biophysics↗

bioRxiv2025

Tau hyperphosphorylation impairs cooperative binding tomicrotubules and perturbs organelle trafficking in neurons

Tau, a neuronal microtubule-associated protein (MAP), organizes the axonal cytoskeleton and regulates intracellular transport. Tau hyperphosphorylation is linked to neurodegeneration in tauopathies including Alzheimers disease. Tau binds microtubules cooperatively to form cohesive envelopes, which are thought to control access to the microtubule lattice and regulate the activity of motor proteins and other microtubule-associated proteins. However, how disease-related perturbations affect tau dynamics and its function as a selective barrier to intracellular transport remains unclear. Using tau phospho-variants in vitro and in live neurons, we show that tau hyperphosphorylation disrupts cooperative microtubule binding and dysregulates lysosome transport. Hyperphosphorylated tau does not form envelopes, distributes more uniformly along the axon, and dissociates faster from microtubules. Tau weakly inhibits KIF5C motility, but strongly inhibits KIF1A. Hyperphosphorylation reduces KIF5C inhibition but increases KIF1A inhibition by decreasing processivity and accelerating detachment. Consistent with these effects, hyperphosphorylated tau alters lysosome transport in neurons. While phospho-resistant tau inhibits processive lysosome motility, hyperphosphorylated tau weakens tau-mediated regulation of lysosome transport, mimicking tau knockout neurons that exhibit enhanced processivity. Altogether, these findings show that hyperphosphorylation disrupts tau envelopes and impairs lysosome trafficking, likely contributing to early defects in degradative pathways that drive neurodegeneration.

biophysics↗

bioRxiv2025

Competition for microtubule lattice spacing between a microtubule expander and compactor.

Microtubules exist in expanded and compacted states, as defined by the lattice spacing of {beta}-tubulin dimers. Changes in lattice spacing has been linked to factors such as GTP-hydrolysis, the binding of microtubule-associated proteins (MAPs), the tubulin code, and microtubule bending. These diverse factors exert opposing molecular driving forces on the microtubule lattice that push lattice spacing towards expanded or compacted states. To better understand how these opposing forces are reconciled, we developed in vitro and cell-based model systems for the competition between a microtubule expander (paclitaxel) and a microtubule compactor (Doublecortin, or DCX). Using an in vitro reconstitution approach, we show that paclitaxel expands microtubules cooperatively. In cells, high concentrations of paclitaxel cause DCX to relocalize to compacted lattices found at concave bends. When the concentration of DCX is increased, however, we find that DCX re-compacts the previously expanded microtubules in vitro. Consistently, high expression levels of DCX prevent its relocalization in paclitaxel treated cells. When the competition between paclitaxel and DCX is "balanced", we observe a complex phenotype: DCX simultaneously localized to both long, straight clusters and concave bends, while other regions on the microtubule network remained DCX-free. We conclude that multiple lattice spacings can coexist in cells. Our results indicate that competition for microtubule lattice spacing is a critical aspect of microtubule physiology.

cell biology↗

bioRxiv2025

Axonal Mechanotransduction Drives Cytoskeletal Responses to Physiological Mechanical Forces

Axons experience strong mechanical forces due to animal movement. While these forces serve as sensory cues in mechanosensory neurons, their impact on other neuron types remains poorly defined. Here, we uncover signaling that controls an axonal cytoskeletal response to external physiological forces and plays a key role in axonal integrity. Live imaging of microtubules at single-polymer resolution in a C. elegans motor neuron reveals local oscillatory movements that fine-tune polymer positioning. Combining cell-specific chemogenetic silencing with targeted degradation alleles to distinguish neuron-intrinsic from extrinsic regulators of these movements, we find that they are driven by muscle contractions and require the mechanosensitive protein Talin, the small GTPase RhoA, and actomyosin activity in the axon. Genetic perturbation of the axons ability to buffer tension by disrupting the spectrin-based membrane-associated skeleton leads to RhoA hyperactivation, actomyosin relocalization to foci at microtubule ends, and converts local oscillations into processive bidirectional movements. This results in large gaps between microtubules, disrupting coverage of the axon and leading to its breakage and degeneration. Notably, hyperpolarizing muscle or degrading components of the mechanotransduction signaling pathway in the axon rescues cytoskeletal defects in spectrin-deficient axons. These results identify mechanisms of an axonal cytoskeletal response to physiological forces and highlight the importance of force-buffering and mechanotransduction signaling for axonal integrity.

neuroscience↗

bioRxiv2024

Huntingtin polyglutamine expansions misdirect axonal transport by perturbing motor and adaptor recruitment

Huntingtons disease (HD) is caused by polyglutamine (polyQ) expansions in huntingtin (HTT). Polyglutamine repeat lengths >35Q lead to neurodegeneration and longer repeats correspond to earlier symptom onset. HTT scaffolds kinesin-1 and dynein to a variety of vesicles and organelles directly and through adaptors. To characterize the effects of HTT polyQ expansions on axonal transport, we tracked BDNF vesicles, mitochondria, and lysosomes in neurons induced from an isogenic set of human stem cell lines with repeat lengths of 30, 45, 65, and 81Q. Mild and intermediate pathogenic polyQ expansions caused increased BDNF motility, while HTT-81Q misdirected BDNF towards the distal tip. In comparison, mitochondria and lysosome transport showed mild defects with polyQHTT. We next examined the effect of polyQHTT in combination with neuroinflammatory stress. Under stress, BDNF cargoes in HTT-30Q neurons were more processive. Stress in HTT-81Q resulted in a stark decrease in the number of BDNF cargoes. However, the few remaining BDNF cargoes displayed more frequent long-range motility in both directions. Under neuroinflammatory stress, lysosomes were more abundant in HTT-81Q neurons, and motile lysosomes moved less processively and had an anterograde bias while lysosomes in HTT-30Q where not strongly affected. To examine how HTT-polyQ expansions altered the motors and adaptors on vesicular cargoes, we isolated BDNF cargoes from neurons and quantified the proteins associated with them. BDNF-endosomes isolated from HTT-81Q neurons associated with 2.5 kinesin-1 and 3.9 HAP1 molecules on average, compared to 1.0 kinesin-1 and 1.0 HAP1 molecule for HTT-30Q neurons. Together, these results show that polyQ expansions in HTT cause aberrant motor and adaptor recruitment to cargoes, resulting in dysregulated transport and responses to neuroinflammatory stress.

cell biology↗

bioRxiv2024

Microtubule perturbations of neurite extension and nucleokinesis in an iPSC-derived model system

In neurons, patterns of different microtubule types are essential for neurite extension and nucleokinesis. Cellular model systems such as rodent primary cultures and induced pluripotent stem cells (iPSC)-derived neurons have provided key insights into how these patterns are created and maintained through the action of microtubule-associated proteins (MAPs), motor proteins, and regulatory enzymes. iPSC-derived models show tremendous promise but lack benchmarking and validation relative to rodent primary cultures. Here we have characterized a recent iPSC-derived model, in which doxycycline-induced expression of Neurogenin-2 drives consistent trans-differentiation into the neuronal state (EBiSC-NEUR1 neurons, referred to as NGN2 neurons below). We developed a suite of open-access, semi-automated methods to measure neurite extension and nucleokinesis of NGN2 neurons, which compare favorably to published data from other models. Then, we challenged NGN2 neurons with a panel of drugs that perturb microtubule physiology. NGN2 neurons extension and nucle-okinesis were significantly perturbed by two microtubule-targeting drugs, namely a taxane (paclitaxel) and a vinca alkaloid (DZ-2384). In contrast, inhibition of microtubule severing (spastazoline) or of deacetylation (trichostatin A) had a limited effect on nucleokinesis only. Our results support the primary importance of microtubule dynamics in neuronal development and demonstrate the power of NGN2 neurons as a model system.

cell biology↗

bioRxiv2023

Doublecortin regulates neuronal migration by editing the tubulin code

Doublecortin (DCX) is a neuronal microtubule-associated protein (MAP) that binds directly to microtubules via two Doublecortin (DC) domains. The DC domains sense the nucleotide state, longitudinal curvature, and protofilament number of the microtubule lattice, indicating a role in the regulation of microtubule structure in neurons. Mutations in DCX cause lissencephaly and subcortical band heterotopia (also known as double-cortex syndrome) due to impaired neuronal migration. To better understand the role of DCX in neuronal migration, we developed a model system based on induced pluripotent stem cells (iPSCs). We used CRISPR/Cas9 to knock-out the Dcx gene in iPSCs and differentiated the cells into cortical neurons. Compared to control neurons, the DCX-KO neurons showed reduced velocities of nuclear movements. The reduced velocities coincided with an increase in the number of neurites early in neuronal development, consistent with a neuronal migration phenotype and previous findings in a DCX-KO mouse model. Neurite branching is regulated by a host of MAPs and other factors, as well as by microtubule polymerization dynamics. However, EB comet dynamics were unchanged in DCX-KO neurons, with similar growth rates, lifetimes, and numbers. Rather, we observed a significant reduction in -tubulin polyglutamylation in DCX-KO neurons. Polyglutamylation levels and neuronal branching were rescued by expression of DCX or of TTLL11, an -tubulin glutamylase. Using U2OS cells as an orthogonal model system, we show that DCX and TTLL11 act synergistically to promote polyglutamylation. Polyglutamylation regulates numerous MAPs, severing enzymes, and molecular motors. Consistently, we observe that lysosomes in DCX-KO neurons show a reduction of their processivity. We propose that the DCX acts as a positive regulator of -tubulin polyglutamylation and restricts neurite branching. Our results indicate an unexpected role for DCX in the homeostasis of the tubulin code.

cell biology↗

bioRxiv2023

Optogenetic control of kinesins -1, -2, -3, and dynein reveals their specific roles in vesicular transport

Each cargo in a cell employs a unique set of motor proteins for its transport. Often multiple types of kinesins are bound to the same cargo. It is puzzling why several types of motors are required for robust transport. To dissect the roles of each type of motor, we developed optogenetic inhibitors of kinesin-1, -2, -3 and dynein. This system allows us to control the activity of the endogenous set of motor proteins that are bound to intracellular cargoes. We examined the effect of optogenetic inhibition of kinesins-1, -2, and -3 and dynein on the transport of early endosomes, late endosomes, and lysosomes. While kinesin-1, kinesin-3, and dynein transport vesicles at all stages of endocytosis, kinesin-2 primarily drives late endosomes and lysosomes. In agreement with previous studies, sustained inhibition of either kinesins or dynein results in reduced motility in both directions. However, transient, optogenetic inhibition of kinesin-1 or dynein causes both early and late endosomes to move more processively by relieving competition with opposing motors. In contrast, optogenetic inhibition of kinesin-2 reduces the motility of late endosomes and lysosomes, and inhibition of kinesin-3 reduces the motility of endosomes and lysosomes. These results suggest that the directionality of transport is likely controlled through regulating kinesin-1 and dynein activity. On vesicles transported by several kinesin and dynein motors, motility can be directed by modulating the activity of a single type of motor on the cargo.

biophysics↗

bioRxiv2022

Tau inhibits the long-range, dynein-mediated motility of early phagosomes

The misregulation of tau, a neuronal microtubule-associated protein, is linked to defective axonal transport and neurodegenerative disease. We reconstituted the motility of isolated phagosomes along microtubules to ask how the sets of motors transporting a cargo determine its motility and response to tau. Using quantitative photobleaching, we find that early phagosomes (EPs) and late phagosomes (LPs) are associated with different sets of kinesin-1, -2, -3, and dynein. While EPs exhibit unidirectional retrograde transport, LPs move bidirectionally. Previously, we found that tau biases LP transport towards the microtubule minus-end. Here, we find that tau strongly inhibits long-range retrograde EP motility. Tau impedes the forces generated by multi-dynein teams and accelerates dynein unbinding under load. Thus, specific cargoes differentially respond to tau, where dynein-complexes on EPs are more sensitive to tau than those on LPs.

biophysics↗

bioRxiv2022

Huntingtin S421 phosphorylation increases kinesin and dynein engagement on early endosomes and lysosomes

Huntingtin (HTT) is a scaffolding protein that recruits motor proteins to vesicular cargoes, enabling it to regulate kinesin-1, dynein, and myosin-VI-dependent transport. To maintain the native stoichiometry of huntingtin with its interacting partners, we used CRISPR/Cas9 to induce a phosphomimetic mutation of the endogenous HTT at S421 (HTT-S421D). Using single particle tracking, optical tweezers, and immunofluorescence, we examined the effects of this mutation on the motility of early endosomes and lysosomes. In HTT-S421D cells, lysosomes exhibit longer displacements and higher processive fractions compared to wild-type (HTT-WT) cells. Kinesins and dyneins exert greater forces on early endosomes and lysosomes in cells expressing HTT-S421D. Additionally, endosomes bind to microtubules faster and are more resistant to detachment under load. The recruitment of kinesins and dyneins to microtubules is enhanced in HTT-S421D cells. In contrast, overexpression of HTT had variable effects on the processivity, displacement, and directional bias of both early endosomes and lysosomes. These data indicate that phosphorylation of the endogenous huntingtin causes early endosomes and lysosomes to move longer distances and more processively by recruiting and activating both kinesin-1 and dynein. Statement of SignificanceThe ubiquitous scaffolding protein huntingtin regulates the recruitment and activity of microtubule motors. Huntingtin phosphorylation at S421 enhances the microtubule binding and force generation of kinesin and dynein on early endosomes and lysosomes. Using optical tweezers to measure the forces exerted on endosomes in CRISPR-engineered cells, we find that a phosphomimetic huntingtin mutation (S421D) enhances both kinesin- and dynein-driven forces on early endosomes and lysosomes. The ability to modulate motor activity on a range of organelles makes huntingtin unique and suggests a significant role for huntingtin in regulating intracellular transport.

biophysics↗

bioRxiv2021

Tau differentially regulates the transport of early endosomes and lysosomes

Microtubule-associated proteins (MAPs) modulate the motility of kinesin and dynein along microtubules to control the transport of vesicles and organelles. The neuronal MAP tau inhibits kinesin-dependent transport. Phosphorylation of tau at tyrosine 18 by fyn kinase results in weakened inhibition of kinesin-1. We examined the motility of early endosomes and lysosomes in cells expressing wild-type (WT) tau and phosphomimetic Y18E tau. Lysosome motility is strongly inhibited by tau. Y18E tau preferentially inhibits lysosomes in the cell periphery, while centrally located lysosomes are less affected. Early endosomes are more sensitive to tau than lysosomes, and are inhibited by both WT and Y18E tau. Our results show that different cargoes have disparate responses to tau, likely governed by the types of kinesin motors driving their transport. In support of this model, kinesin-1 and -3 are strongly inhibited by tau while kinesin-2 and dynein are less affected. In contrast to kinesin-1, we find that kinesin-3 is strongly inhibited by phosphorylated tau.

biophysics↗

bioRxiv2021

The Pathogenic R5L Mutation Disrupts Formation of Tau Complexes on the Microtubule by Altering Local N-Terminal Structure

The microtubule-associated protein (MAP) Tau is an intrinsically disordered protein (IDP) primarily expressed in axons, where it functions to regulate microtubule dynamics, modulate motor protein motility, and participate in signaling cascades. Tau misregulation and point mutations are linked to neurodegenerative diseases, including Progressive Supranuclear Palsy (PSP), Picks Disease and Alzheimers disease. Many disease-associated mutations in Tau occur in the C-terminal microtubule-binding domain of the protein. Effects of C-terminal mutations in Tau have led to the widely accepted disease-state theory that missense mutations in Tau reduce microtubule-binding affinity or increase Tau propensity to aggregate. Here, we investigate the effect of an N-terminal disease-associated mutation in Tau, R5L, on Tau-microtubule interactions using an in vitro reconstituted system. Contrary to the canonical disease-state theory, we determine the R5L mutation does not reduce Tau affinity for the microtubule using Total Internal Reflection Fluorescence (TIRF) Microscopy. Rather, the R5L mutation decreases the ability of Tau to form larger order complexes, or Tau patches, at high concentrations of Tau. Using Nuclear Magnetic Resonance (NMR), we show that the R5L mutation results in a local structural change that reduces interactions of the projection domain in the presence of microtubules. Altogether, these results challenge both the current paradigm of how mutations in Tau lead to disease and the role of the projection domain in modulating Tau behavior on the microtubule surface. Significance StatementThe microtubule-associated protein Tau is strongly linked to a number of neurological diseases. Disease onset is typically associated with weakened interaction with the microtubule, but this widely accepted model is based on hyperphosphorylation or mutations within the C-terminal microtubule-binding domain of Tau. Here, we find an N-terminal disease-associated mutation in Tau, R5L, does not reduce Tau affinity for microtubules, but instead modifies the N-terminal structure, altering Taus behavior and ability to condense on the microtubule surface. Our findings challenge the current paradigms of both how mutations in Tau lead to disease and the functional role of the N-terminal region of Tau.

biophysics↗

bioRxiv2021

The mechanochemical origins of the microtubule sliding motility within the kinesin-5 domain organization

The conserved kinesin-5 bipolar tetrameric motors slide apart microtubules during mitotic spindle assembly and elongation. Kinesin-5 bipolar organization originates from its conserved tetrameric helical minifilament, which position the C-terminal tail domains of two subunits near the N-terminal motor domains of two anti-parallel subunits (Scholey et al, 2014). This unique tetrameric structure enables kinesin-5 to simultaneously engage two microtubules and transmit forces between them, and for multiple kinesin-5 motors to organize via tail to motor interactions during microtubule sliding (Bodrug et al, 2020). Here, we show how these two structural adaptations, the kinesin-5 tail-motor domain interactions and the length of the tetrameric minifilament, determine critical aspects of kinesin-5 motility and sliding mechanisms. An x-ray structure of the 34-nm kinesin-5 minifilament reveals how the dual dimeric N-terminal coiled-coils emerge from the tetrameric central bundle. Using this structure, we generated active bipolar mini-tetrameric motors from Drosophila and human orthologs, which are half the length of native kinesin-5. Using single-molecule motility assays, we show that kinesin-5 tail domains promote mini-tetramers static pauses that punctuate processive motility. During such pauses, kinesin-5 mini-tetramers form multi-motor clusters mediated via tail to motor domain cross-interactions. These clusters undergo slow and highly processive motility and accumulate at microtubule plus-ends. In contrast to native kinesin-5, mini-tetramers require tail domains to initiate microtubule crosslinking. Although mini-tetramers are highly strained in initially aligning microtubules, they slide microtubules more efficiently than native kinesin-5, due to their decreased minifilament flexibility. Our studies reveal that the conserved kinesin-5 motor-tail mediated clustering and the length of the tetrameric minifilament are key features for sliding motility and are critical in organizing microtubules during mitotic spindle assembly and elongation.

cell biology↗

bioRxiv2019

Dynamic crosslinking of the actin cytoskeleton governs cell mechanics

Cells precisely control their mechanical properties to organize and differentiate into tissues. The architecture and connectivity of cytoskeletal filaments changes in response to mechanical and biochemical cues, allowing the cell to rapidly tune its mechanics from highly-crosslinked, elastic networks to weakly-crosslinked viscous networks. While the role of actin crosslinking in controlling actin network mechanics is well-characterized in purified actin networks, its mechanical role in the cytoplasm of living cells remains unknown. Here, we probe the frequency-dependent intracellular viscoelastic properties of living cells using multifrequency excitation and in situ optical trap calibration. At long timescales in the intracellular environment, we observe that the cytoskeleton becomes fluid-like. The mechanics are well-captured by a model in which actin filaments are dynamically connected by a single dominant crosslinker. A disease-causing point mutation (K255E) of the actin crosslinker -actinin 4 (ACTN4) causes its binding kinetics to be insensitive to tension. Under normal conditions, the viscoelastic properties of wild type (WT) and K255E+/- cells are similar. However, when tension is reduced through myosin II inhibition, WT cells relax 3x faster to the fluid-like regime while K255E+/- cells are not affected. These results indicate that dynamic actin crosslinking enables the cytoplasm to flow at long timescales.

biophysics↗

bioRxiv2019

MAP7 recruits kinesin-1 to microtubules to direct organelle transport

Microtubule-associated proteins (MAPs) play well-characterized roles in regulating microtubule polymerization, dynamics, and organization. In addition, MAPs control trans-port along microtubules by regulating the motility of kinesin and dynein. MAP7 (ensconsin, E-MAP-115) is a ubiquitous MAP that organizes the microtubule cytoskeleton in mitosis and neuronal branching. MAP7 also promotes the interaction of kinesin-1 with microtubules. We expressed and purified full-length kinesin-1 and MAP7 in Sf9 cells. In single-molecule motiity assays, MAP7 recruits kinesin-1 to microtubules, increasing the frequency of both diffusive and processive runs. Optical trapping assays on beads transported by single and teams of kinesin-1 motors indicate that MAP7 increases the relative binding rate of kinesin-1 and the number of motors simultaneously engaged in ensembles. To examine the role of MAP7 in regulating bidirectional transport, we isolated late phagosomes along with their native set of kinesin-1, kinesin-2, and dynein motors. Bidirectional cargoes exhibit a clear shift towards plus-end directed motility on MAP7-decorated microtubules due to increased forces exerted by kinesin teams. Collectively, our results indicate that MAP7 enhances kinesin-1 recruitment to microtubules and targets organelle transport to the plus end.

biophysics↗

bioRxiv2017

Tau directs intracellular trafficking by regulating the forces exerted by kinesin and dynein teams

Organelles, proteins, and mRNA are transported bidirectionally along microtubules by plus-end directed kinesin and minus-end directed dynein motors. Microtubules are decorated by microtubule-associated proteins (MAPs) that organize the cytoskeleton, regulate microtubule dynamics and modulate the interaction between motor proteins and microtubules to direct intracellular transport. Tau is a neuronal MAP that stabilizes axonal microtubules and crosslinks them into bundles. Dysregulation of tau leads to a range of neurodegenerative diseases known as tauopathies including Alzheimers disease (AD). Tau reduces the processivity of kinesin and dynein by acting as an obstacle on the microtubule. Single-molecule assays indicate that kinesin-1 is more strongly inhibited than kinesin-2 or dynein, suggesting tau might act to spatially modulate the activity of specific motors. To investigate the role of tau in regulating bidirectional transport, we isolated phagosomes driven by kinesin-1, kinesin-2, and dynein and reconstituted their motility along microtubules. We find that tau biases bidirectional motility towards the microtubule minus-end in a dose-dependent manner. Optical trapping measurements show that tau increases the magnitude and frequency of forces exerted by dynein through inhibiting opposing kinesin motors. Mathematical modeling indicates that tau controls the directional bias of intracellular cargoes through differentially tuning the processivity of kinesin-1, kinesin-2, and dynein. Taken together, these results demonstrate that tau modulates motility in a motor-specific manner to direct intracellular transport, and suggests that dysregulation of tau might contribute to neurodegeneration by disrupting the balance of plus- and minus-end directed transport.\n\nSynopsis and Graphical Table of ContentsWe isolated endogenous cargoes, along with a complement of kinesin-1, kinesin-2, and dynein motors, and reconstituted their bidirectional motility in vitro. We find that tau, a microtubule-associated protein that stabilizes microtubules in neuronal axons, directs bidirectional cargoes towards the microtubule minus end by tuning the balance of forces exerted by kinesin and dynein teams. These results suggest a general mechanism for regulating the transport of intracellular cargoes through modulating the relative activity of opposing motor teams.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC=\"FIGDIR/small/173609_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (31K):\norg.highwire.dtl.DTLVardef@8ba4e0org.highwire.dtl.DTLVardef@1f5945corg.highwire.dtl.DTLVardef@1df2f47org.highwire.dtl.DTLVardef@198f5d8_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2017

Endocytosis caused by liquid-liquid phase separation of proteins

Membrane invagination and vesicle formation are key steps in endocytosis and cellular trafficking. Here, we show that endocytic coat proteins with prion-like domains (PLDs) form hemispherical puncta in the budding yeast, S. cerevisiae. These puncta have the hallmarks of biomolecular condensates and enable membrane remodeling to drive actin-independent endocytosis. The puncta, which we refer to as endocytic condensates, form and dissolve reversibly in response to changes in temperature and solution conditions. The condensates are organized around dynamic protein-protein interaction networks, which involve interactions among PLDs with high glutamine contents. The endocytic coat protein Sla1 is at the hub of the protein-protein interaction network. Using active rheology, we indirectly characterized the material properties of endocytic condensates. These experiments show that endocytic condensates are viscoelastic materials and allow us to estimate the interfacial tension between endocytic condensates and their surroundings. We then adapt the physics of contact mechanics, specifically the contact theory of Hertz, to develop a quantitative framework for describing how interfacial tensions among condensates, the membrane, and the cytosol can deform the plasma membrane to enable actin independent endocytosis.

cell biology↗