Ben Goult | University of Liverpool (original) (raw)

Papers by Ben Goult

Open Biology, 2024

Misprocessing of amyloid precursor protein (APP) is one of the major causes of Alzheimer’s diseas... more Misprocessing of amyloid precursor protein (APP) is one of the major
causes of Alzheimer’s disease. APP comprises a large extracellular region,
a single transmembrane helix and a short cytoplasmic tail containing
an NPxY motif (normally referred to as the YENPTY motif). Talins are
synaptic scaffold proteins that connect the cytoskeletal machinery to the
plasma membrane via binding NPxY motifs in the cytoplasmic tail of
integrins. Here, we report the crystal structure of an APP/talin1 complex
identifying a new way to couple the cytoskeletal machinery to synaptic
sites through APP. Proximity ligation assay (PLA) confirmed the close
proximity of talin1 and APP in primary neurons, and talin1 depletion
had a dramatic effect on APP processing in cells. Structural modelling
reveals APP might form an extracellular meshwork that mechanically
couples the cytoskeletons of the pre- and post-synaptic compartments.
We propose APP processing represents a mechanical signalling pathway
whereby under tension, the cleavage sites in APP have varying accessibility
to cleavage by secretases. This leads us to propose a new hypothesis
for Alzheimer’s, where misregulated APP dynamics result in loss of the
mechanical integrity of the synapse, corruption and loss of mechanical
binary data, and excessive generation of toxic plaque-forming Aβ42
peptide.

PLOS genetics, Apr 25, 2024

bioRxiv (Cold Spring Harbor Laboratory), Mar 6, 2024

Current opinion in cell biology, Apr 1, 2024

Cells sense and respond to subtle changes in their physicality, and via a myriad of different mec... more Cells sense and respond to subtle changes in their physicality, and via a myriad of different mechanosensitive processes, convert these physical cues into chemical and biochemical signals. This process, called mechanotransduction, is possible due to a highly sophisticated machinery within cells. One mechanism by which this can occur is via the stretching of mechanosensitive proteins. Stretching proteins that contain force-dependent regions results in altered geometry and dimensions of the connections, as well as differential spatial organization of signals bound to the stretched protein. The purpose of this mini-review is to discuss some of the intense recent activity in this area of mechanobiology that strives to understand how protein stretching can influence signaling outputs and cellular responses.

PLOS Computational Biology, Oct 5, 2023

Cells interact with the extracellular matrix (ECM) via cell-ECM adhesions. These physical interac... more Cells interact with the extracellular matrix (ECM) via cell-ECM adhesions. These physical interactions are transduced into biochemical signals inside the cell which influence cell behaviour. Although cell-ECM interactions have been studied extensively, it is not completely understood how immature (nascent) adhesions develop into mature (focal) adhesions and how mechanical forces influence this process. Given the small size, dynamic nature and short lifetimes of nascent adhesions, studying them using conventional microscopic and experimental techniques is challenging. Computational modelling provides a valuable resource for simulating and exploring various "what if?" scenarios in silico and identifying key molecular components and mechanisms for further investigation. Here, we present a simplified mechano-chemical model based on ordinary differential equations with three major proteins involved in adhesions: integrins, talin and vinculin. Additionally, we incorporate a hypothetical signal molecule that influences adhesion (dis)assembly rates. We find that assembly and disassembly rates need to vary dynamically to limit maturation of nascent adhesions. The model predicts biphasic variation of actin retrograde velocity and maturation fraction with substrate stiffness, with maturation fractions between 18-35%, optimal stiffness of *1 pN/nm, and a mechanosensitive range of 1-100 pN/nm, all corresponding to key experimental findings. Sensitivity analyses show robustness of outcomes to small changes in parameter values, allowing model tuning to reflect specific cell types and signaling cascades. The model proposes that signal-dependent disassembly rate variations play an underappreciated role in maturation fraction regulation, which should be investigated further. We also provide predictions on the changes in traction force generation under increased/decreased vinculin concentrations, complementing previous vinculin overexpression/knockout experiments in different cell types. In summary, this work proposes a model framework to robustly simulate the mechanochemical processes underlying adhesion maturation and maintenance, thereby enhancing our fundamental knowledge of cell-ECM interactions.

The cAMP-dependent protein kinase (Protein Kinase A; PKA) is a ubiquitous, promiscuous kinase who... more The cAMP-dependent protein kinase (Protein Kinase A; PKA) is a ubiquitous, promiscuous kinase whose activity is focused and specified through subcellular localization mediated by A-kinase anchoring proteins (AKAPs). PKA has complex roles as both an effector and a regulator of integrin-mediated cell adhesion to the extracellular matrix (ECM). Recent observations demonstrate that PKA is an active component of focal adhesions (FA), intracellular complexes coupling ECM-bound integrins to the actin cytoskeleton, suggesting the existence of one or more FA AKAPs. Using a combination of a promiscuous biotin ligase fused to PKA type-IIα regulatory (RIIα) subunits and subcellular fractionation, we identify the archetypal FA protein talin1 as an AKAP. Talin is a large, mechanosensitive scaffold that directly links integrins to actin filaments and promotes FA assembly by recruiting additional components in a force-dependent manner. The rod region of talin1 consists of 62 α-helices bundled into ...

Science Advances, 2024

Tissue mechanical properties are determined mainly by the extracellular matrix (ECM) and actively... more Tissue mechanical properties are determined mainly by the extracellular matrix (ECM) and actively maintained by resident cells. Despite its broad importance to biology and medicine, tissue mechanical homeostasis remains poorly understood. To explore cell-mediated control of tissue stiffness, we developed mutations in the mechano- sensitive protein talin 1 to alter cellular sensing of ECM. Mutation of a mechanosensitive site between talin 1 rod- domain helix bundles R1 and R2 increased cell spreading and tension exertion on compliant substrates. These mutations promote binding of the ARP2/3 complex subunit ARPC5L, which mediates the change in substrate stiff- ness sensing. Ascending aortas from mice bearing these mutations showed less fibrillar collagen, reduced axial stiffness, and lower rupture pressure. Together, these results demonstrate that cellular stiffness sensing contrib- utes to ECM mechanics, directly supporting the mechanical homeostasis hypothesis and identifying a mechano- sensitive interaction within talin that contributes to this mechanism.

PLoS Computational Biology, 2024

Vinculin binds to specific sites of mechanically unfolded talin rod domains to reinforce the coup... more Vinculin binds to specific sites of mechanically unfolded talin rod domains to reinforce the coupling of the cell's exterior to its force generation machinery. Force-dependent vinculintalin complexation and dissociation was previously observed as contraction or extension of the unfolded talin domains respectively using magnetic tweezers. However, the structural mechanism underlying vinculin recognition of unfolded vinculin binding sites (VBSs) in talin remains unknown. Using molecular dynamics simulations, we demonstrate that a VBS dynamically refolds under force, and that vinculin can recognize and bind to partially unfolded VBS states. Vinculin binding enables refolding of the mechanically strained VBS and stabilizes its folded α-helical conformation, providing resistance against mechanical stress. Together, these results provide an understanding of a recognition mechanism of proteins unfolded by force and insight into the initial moments of how vinculin binds unfolded talin rod domains during the assembly of this mechanosensing meshwork.

Journal of Cell Biology, 2024

We previously identified talin rod domain-containing protein 1 (TLNRD1) as a potent actin-bundlin... more We previously identified talin rod domain-containing protein 1 (TLNRD1) as a potent actin-bundling protein in vitro. Here, we report that TLNRD1 is expressed in the vasculature in vivo. Its depletion leads to vascular abnormalities in vivo and modulation of endothelial cell monolayer integrity in vitro. We demonstrate that TLNRD1 is a component of the cerebral cavernous malformations (CCM) complex through its direct interaction with CCM2, which is mediated by a hydrophobic C-terminal helix in CCM2 that attaches to a hydrophobic groove on the four-helix domain of TLNRD1. Disruption of this binding interface leads to CCM2 and TLNRD1 accumulation in the nucleus and actin fibers. Our findings indicate that CCM2 controls TLNRD1 localization to the cytoplasm and inhibits its actin-bundling activity and that the CCM2-TLNRD1 interaction impacts endothelial actin stress fiber and focal adhesion formation. Based on these results, we propose a new pathway by which the CCM complex modulates the actin cytoskeleton and vascular integrity.

Nature Communications, 2024

Force transmission through adherens junctions (AJs) is crucial for multicellular organization, wo... more Force transmission through adherens junctions (AJs) is crucial for multicellular organization, wound healing and tissue regeneration. Recent studies shed light on the molecular mechanisms of mechanotransduction at the AJs. However, the canonical model fails to explain force transmission when essential proteins of the mechanotransduction module are mutated or missing. Here, we demonstrate that, in absence of α-catenin, β-catenin can directly and func- tionally interact with vinculin in its open conformation, bearing physiological forces. Furthermore, we found that β-catenin can prevent vinculin auto- inhibition in the presence of α-catenin by occupying vinculin ́s head-tail interaction site, thus preserving force transmission capability. Taken together, our findings suggest a multi-step force transmission process at AJs, where α- catenin and β-catenin can alternatively and cooperatively interact with vin- culin. This can explain the graded responses needed to maintain tissue mechanical homeostasis and, importantly, unveils a force-bearing mechanism involving β-catenin and extended vinculin that can potentially explain the underlying process enabling collective invasion of metastatic cells lacking α-catenin.

PNAS, 2024

There is increasing interest in the ability of proteins involved in cell adhesion to convert mech... more There is increasing interest in the ability of proteins involved in cell adhesion to convert mechanical force into altered biochemistry. Cellular signal transduction is most often conducted through multiprotein scaffolds that consolidate, localize, and specify signaling inputs and outputs. This report bridges these fields by identifying an interaction between talin, a mechanosensitive adhesion protein, and PKA, a pleiotropic kinase with myriad cellular targets. Together, these observations form the foundation for a mechanotransduction pathway that utilizes forcedependent changes in protein conformation to establish a solid-state signaling complex well positioned to couple cellular tension to cellular communication.

Current Opinion in Cell Biology, 2024

PLoS Genetics, 2024

Cell adhesion requires linkage of transmembrane receptors to the cytoskeleton through intermediar... more Cell adhesion requires linkage of transmembrane receptors to the cytoskeleton through intermediary linker proteins. Integrin-based adhesion to the extracellular matrix (ECM) involves large adhesion complexes that contain multiple cytoskeletal adapters that connect to the actin cytoskeleton. Many of these adapters, including the essential cytoskeletal linker Talin, have been shown to contain multiple actin-binding sites (ABSs) within a single protein. To investigate the possible role of having such a variety of ways of linking integrins to the cytoskeleton, we generated mutations in multiple actin binding sites in Drosophila talin. Using this approach, we have been able to show that different actin-binding sites in talin have both unique and complementary roles in integrin-mediated adhesion. Specifically, mutations in either the C-terminal ABS3 or the centrally located ABS2 result in lethality showing that they have unique and non-redundant function in some contexts. On the other hand, flies simultaneously expressing both the ABS2 and ABS3 mutants exhibit a milder phenotype than either mutant by itself, suggesting overlap in function in other contexts. Detailed phenotypic analysis of ABS mutants elucidated the unique roles of the talin ABSs during embryonic development as well as provided support for the hypothesis that talin acts as a dimer in in vivo contexts. Overall, our work highlights how the ability of adhesion complexes to link to the cytoskeleton in multiple ways provides redundancy, and consequently robustness, but also allows a capacity for functional specialization.

bioRxiv, 2024

Misprocessing of Amyloid Precursor Protein (APP) is one of the major causes of Alzheimer's diseas... more Misprocessing of Amyloid Precursor Protein (APP) is one of the major causes of Alzheimer's disease. APP is a transmembrane protein comprising a large extracellular region, a single transmembrane helix and a short cytoplasmic tail containing an NPxY motif (normally referred to as the YENPTY motif). Talin is a synaptic scaffold protein that connects the cytoskeletal machinery to the plasma membrane via binding to one of two highly conserved NPxY motifs in the cytoplasmic tail of integrin transmembrane receptors. Here we report the crystal structure of an APP/talin complex identifying a new way to couple the cytoskeletal machinery to synaptic sites via APP. Structural modelling reveals that APP forms an extracellular meshwork that mechanically couples the cytoskeletal meshworks of both the pre-, and post-synaptic compartments. In this context, we propose APP processing as a mechanical signalling pathway with similarities to Notch signalling, whereby the cleavage sites in APP represent mechanical sensors, with varying accessibility to cleavage by secretases. During synaptogenesis in healthy neurons, the APP/talin linkage would provide an exquisite mechanical coupling between synapses, with tightly controlled APP processing providing instructions to maintain this synchrony. Furthermore, APP directly coupling to the binary switches in talin indicates a role for APP in mechanical memory storage as postulated by the MeshCODE theory. The implication that APP is a regulator of mechanical signalling leads to a new hypothesis for Alzheimer's disease, where mis-regulation of APP dynamics results in loss of mechanical integrity of the synapse, corruption and loss of mechanical binary data, and excessive generation of the toxic plaque-forming Aβ42 peptide. In support of this model, we show that talin1 depletion has a dramatic effect on APP processing in cells. Much needs to be done to experimentally validate this idea, but we present here a novel theory of Alzheimer's Disease with a role for APP in the mechanically coded binary information storage in the synapse, which identifies a potential novel therapeutic strategy for treating Alzheimer's Disease.

Nature Nanotechnology, 2023

Extreme energy-dissipating materials are essential for a range of applications. The military and ... more Extreme energy-dissipating materials are essential for a range of applications. The military and police force require ballistic armour to ensure the safety of their personnel, while the aerospace industry requires materials that enable the capture, preservation and study of hypervelocity projectiles. However, current industry standards display at least one inherent limitation, such as weight, breathability, stiffness, durability and failure to preserve captured projectiles. To resolve these limitations, we have turned to nature, using proteins that have evolved over millennia to enable effective energy dissipation. Specifically, a recombinant form of the mechanosensitive protein talin was incorporated into a monomeric unit and crosslinked, resulting in a talin shock-absorbing material (TSAM). When subjected to 1.5 km s −1 supersonic shots, TSAMs were shown to absorb the impact and capture and preserve the projectile.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature