Biology subjects

Brockman, J. M.

Publications and source records attributed to Brockman, J. M..

5 recordsLinked to original sources

bioRxiv2026

Substrate mediated mechanical forces enable optimal kinetic proofreading by T-cell receptors

T-cells use molecular reactions with nonequilibrium error correction, i.e., proofreading, to discriminate between nearly identical antigens with high specificity and sensitivity. These receptor binding events are known to be force sensitive, yet traditional schemes of proofreading focus on reaction kinetics alone and do not consider the role of force dependent catch/slip bond behavior or interactions with mechanically engaged coreceptors such as adhesion molecules. To address this, we propose a minimal framework for proofreading of ligand discrimination by T-cell receptors (TCRs) that uses endogenous TCR mechanosensation and substrate-mediated mechanical interactions with adhesive proteins (load sharing) to improve recognition fidelity. We leverage the catch bond behavior of cognate antigens to delay decision making and amplify TCR signaling while discarding noncognate slip bond ligands in the presence of a force. By integrating our model with existing structural and molecular data, we show that substrate mechanics regulates the transmission of active cytoskeletal forces through a molecular clutch and controls the energization of bound TCRs needed for optimal proofreading. Our work demonstrates how mechanical forces and substrate properties can augment kinetic proofreading in T-cells, suggesting biomaterial design strategies for immunotherapies that tune the mechanical microenvironment of T-cells to achieve high fidelity TCR-ligand discrimination, antigen recognition, and activation.

biophysics↗

bioRxiv2025

Prediction of Piconewton Receptor Tension Images using Deep Learning

Piconewton (pN) receptor forces govern many biological processes, but measuring these forces remains challenging. Molecular tension probes (MTPs) provide a sensitive means to measure pN cellular forces via fluorescence microscopy; however, MTPs are challenging to use and only forces transmitted through the probes are reported, complicating their use in heterogenous environments. Here, we present Tension Deep Learning (TensionDL), which leverages convolutional neural networks and image-to-image translation to predict pN receptor tension maps from images of cell morphology and the force-transducing protein vinculin. We validate the accuracy of TensionDL at the subcellular and cellular scales, demonstrate model accuracy across different substrate stiffnesses and cell types, and leverage TensionDL to make semi-quantitative predictions of cell mechanical output. Finally, TensionDL enables long-term mapping of pN receptor tension and infers tension distributions in heterogeneous environments in which some forces are not transduced through MTPs.

bioengineering↗

bioRxiv2025

Chimeric Antigen Receptors Transmit Co-stimulatory Domain Dependent Piconewton Forces to their Target

Chimeric antigen receptor (CAR) T cells promote tumor-specific cytotoxicity through engagement of a recombinant, synthetic receptor with target ligands expressed on cancer cells. Native T cells are mechanically active, both transmitting and sensing forces exceeding 19 piconewtons (pN) via transmembrane receptors, including the T cell receptor (TCR). Emerging evidence implicates mechanoactivity in CAR T cell biology, but CAR-transmitted T cell forces have not been directly measured. Here, we utilize DNA-based molecular tension probes (MTPs) conjugated to CAR target ligands, providing evidence of actin-polymerization dependent forces exceeding 4.7-19 pN borne by the CAR. We demonstrate force transmission by three clinically relevant CARs (CD123, CD33, and CD19), suggesting that these forces are generalizable across CAR targets and constructs. Additionally, we identify intracellular co-stimulatory domains as the main determinants of CAR-mediated forces, because first-generation CARs lacking co-stimulatory domains do not transmit measurable forces to their ligand. Finally, we demonstrate that CAR forces temporally precede Ca2+ signaling and are spatially correlated with phosphorylation of classical TCR-signaling machinery, indicating a link between CAR T cell forces and early biochemical signaling. Our study introduces CAR-mediated mechanobiology as a key correlate of early CAR T cell activation events. Significance StatementChimeric antigen receptor (CAR) T cell therapies have revolutionized treatment for several hematological malignancies. CARs are recombinant receptors containing domains derived from the T cell receptor complex machinery and other co-stimulatory proteins. Mechanical forces are believed to be important in T cell activation and antigen recognition. The role of mechanobiology in CAR T cell immunotherapy remains poorly understood. Here, using DNA-based molecular tension probes conjugated to CAR ligands, we provide direct evidence that CARs bear actin polymerization-dependent piconewton forces during antigen engagement that precede early signaling events. These forces depend on CAR co-stimulatory domains, and first-generation CARs lacking these domains fail to transmit detectable force. These findings suggest mechanobiology may be a key, tunable parameter for next-generation CAR T cells.

immunology↗

bioRxiv2025

Human progenitor T-cell differentiation regulated by the mechanical resistance of thymus-mimetic extracellular matrices

Therapeutic T-cell engineering ex vivo from human hematopoietic stem cells (HSCs) focuses on recapitulating notch1-signaling and 4{beta}1-integrin-mediated adhesion within the thymic niche with supportive stromal cell feeder-layers or surface-immobilized recombinant protein-based engineered thymic niches (ETNs). The relevant Notch1-DLL-4 and 4{beta}1-integrin-VCAM-1 interactions are known to respond to mechanical forces that regulate their bond dissociation behaviors and downstream signal transduction, yet manipulating the mechanosensitive features of these key receptor-ligand interactions in thymopoiesis has been largely ignored in current ETN designs. Here, we demonstrate that human T-cell development from cord blood-derived CD34+ HSCs is regulated via molecular cooperativity in notch1 and integrin-mediated mechanotransduction. Mechanically confining interpenetrating network (IPN) hydrogel-based 3D cell culture comprised of collagen type I and alginate polysaccharides functionalized with DLL-4 and VCAM-1 is used as a model viscoelastic 3D ETN to manipulate human progenitor (pro)T-cell differentiation. This ETN enables orthogonal control of the mechanical and biomolecular features of the thymic niche, including thymopoietic ligand density, modulus, and viscoelastic properties (e.g., stress relaxation kinetics). We identify that soft, viscous matrices that enhance activation of the notch1-pathway, and subsequently notch1 intracellular domain (NICD) nuclear import sustain the T-cell development gene regulatory network during proT-cell differentiation. Conversely, stiff, elastic matrices inhibit HSC commitment to the T-lineage, and rather promotes Myeloid-cell differentiation. Our observations indicate mechanical reciprocity in signaling pathways indispensable to thymopoiesis, and highlights extracellular matrix mechanics as a variable in controlling hematopoietic stem cell fate decisions.

bioengineering↗

bioRxiv2022

Enhancing adoptive T cell therapy with synergistic host immune engagement promotes long-term protection against solid tumors

Adoptive T cell therapy provides the T cell pool needed for immediate tumor debulking, but the infused T cells generally have a narrow repertoire for antigen recognition and limited ability for long-term protection. Here, we present a biomaterial platform that enhances adoptive T cell therapy by synergistically engaging the host immune system via in-situ antigen-free vaccination. T cells alone loaded into these localized cell depots provided significantly better control of subcutaneous B16-F10 tumors than T cells delivered through direct peritumoral injection or intravenous infusion. The anti-tumor response was significantly enhanced when T cell delivery was combined with biomaterial-driven accumulation and activation of host immune cells, as this prolonged the activation state of the delivered T cells, minimized host T cell exhaustion, and enabled long-term tumor control. This integrated approach provides both immediate tumor debulking and long-term protection against solid tumors, including against tumor antigen escape.

immunology↗