Optoregulated force application to cellular receptors using molecular motors

AbstractProgress in our understanding of mechanotransduction events requires noninvasive methods for the manipulation of forces at molecular scale in physiological environments. Inspired by cellular mechanisms for force application (i.e. motor proteins pulling on cytoskeletal fibers), we present a unique molecular machine that can apply forces at cell-matrix and cell-cell junctions using light as an energy source. The key actuator is a light-driven rotatory molecular motor linked to polymer chains, which is intercalated between a membrane receptor and an engineered biointerface. The light-driven actuation of the molecular motor is converted in mechanical twisting of the entangled polymer chains, which will in turn effectively “pull” on engaged cell membrane receptors (e.g., integrins, T cell receptors) within the illuminated area. Applied forces have physiologically-relevant magnitude and occur at time scales within the relevant ranges for mechanotransduction at cell-friendly exposure conditions, as demonstrated in force-dependent focal adhesion maturation and T cell activation experiments. Our results reveal the potential of nanomotors for the manipulation of living cells at the molecular scale and demonstrate a functionality which at the moment cannot be achieved by other technologies for force application.

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Optoregulated force application to cellular receptors using molecular motors

10.1101/2020.03.31.015198 ◽

2020 ◽

Author(s):

Yijun Zheng ◽

Mitchell K.L. Han ◽

Renping Zhao ◽

Johanna Blass ◽

Jingnan Zhang ◽

...

Keyword(s):

T Cell Activation ◽

Molecular Motors ◽

Cell Activation ◽

Molecular Motor ◽

Membrane Receptors ◽

Polymer Chains ◽

Cell Junctions ◽

Cellular Mechanisms ◽

Force Application ◽

Applied Forces

AbstractMechanotransduction events in physiological environments are difficult to investigate, in part due to the lack of experimental tools to apply forces to mechanosensitive receptors remotely. Inspired by cellular mechanisms for force application (i.e. motor proteins pulling on cytoskeletal fibers), here we present a unique molecular machine that can apply forces at cell-matrix and cell-cell junctions using light as an energy source. The key actuator is a light-driven rotatory molecular motor linked to polymer chains, which is intercalated between a membrane receptor and an engineered biointerface. The light-driven actuation of the molecular motor is converted in mechanical twisting of the polymer chains, which will in turn effectively “pulls” on engaged cell membrane receptors (integrins, cadherins…) within the illuminated area. Applied forces have the adequate magnitude and occur at time scales within the relevant ranges for mechanotransduction at cell-friendly exposure conditions, as demonstrated in forcedependent focal adhesion maturation and T cell activation experiments. Our results reveal the potential of nanomotors for the manipulation of living cells at the molecular scale and demonstrate, for the first time, a functionality which at the moment cannot be achieved by any other means.

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Role of Diacylglycerol Kinase α in the Attenuation of Receptor Signaling

The Journal of Cell Biology ◽

10.1083/jcb.153.1.207 ◽

2001 ◽

Vol 153 (1) ◽

pp. 207-220 ◽

Cited By ~ 108

Author(s):

Miguel Angel Sanjuán ◽

David R. Jones ◽

Manuel Izquierdo ◽

Isabel Mérida

Keyword(s):

Plasma Membrane ◽

T Cell ◽

Phosphatidic Acid ◽

T Cell Activation ◽

Cell Activation ◽

Active Form ◽

Diacylglycerol Kinase ◽

Membrane Receptors ◽

Type I ◽

Cell Responses

Diacylglycerol kinase (DGK) is suggested to attenuate diacylglycerol-induced cell responses through the phosphorylation of this second messenger to phosphatidic acid. Here, we show that DGKα, an isoform highly expressed in T lymphocytes, translocates from cytosol to the plasma membrane in response to two different receptors known to elicit T cell activation responses: an ectopically expressed muscarinic type I receptor and the endogenous T cell receptor. Translocation in response to receptor stimulation is rapid, transient, and requires calcium and tyrosine kinase activation. DGKα-mediated phosphatidic acid generation allows dissociation of the enzyme from the plasma membrane and return to the cytosol, as demonstrated using a pharmacological inhibitor and a catalytically inactive version of the enzyme. The NH2-terminal domain of the protein is shown to be responsible for receptor-induced translocation and phosphatidic acid–mediated membrane dissociation. After examining induction of the T cell activation marker CD69 in cells expressing a constitutively active form of the enzyme, we present evidence of the negative regulation that DGKα exerts on diacylglycerol-derived cell responses. This study is the first to describe DGKα as an integral component of the signaling cascades that link plasma membrane receptors to nuclear responses.

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Folding for the Immune Synapse: CCT Chaperonin and the Cytoskeleton

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.658460 ◽

2021 ◽

Vol 9 ◽

Author(s):

Noa Beatriz Martín-Cófreces ◽

José María Valpuesta ◽

Francisco Sánchez-Madrid

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

De Novo ◽

Mrna Translation ◽

Membrane Receptors ◽

Microtubule Network ◽

Activated T Cells ◽

Immune Synapse

Lymphocytes rearrange their shape, membrane receptors and organelles during cognate contacts with antigen-presenting cells (APCs). Activation of T cells by APCs through pMHC-TCR/CD3 interaction (peptide-major histocompatibility complex-T cell receptor/CD3 complexes) involves different steps that lead to the reorganization of the cytoskeleton and organelles and, eventually, activation of nuclear factors allowing transcription and ultimately, replication and cell division. Both the positioning of the lymphocyte centrosome in close proximity to the APC and the nucleation of a dense microtubule network beneath the plasma membrane from the centrosome support the T cell’s intracellular polarity. Signaling from the TCR is facilitated by this traffic, which constitutes an important pathway for regulation of T cell activation. The coordinated enrichment upon T cell stimulation of the chaperonin CCT (chaperonin-containing tailless complex polypeptide 1; also termed TRiC) and tubulins at the centrosome area support polarized tubulin polymerization and T cell activation. The proteasome is also enriched in the centrosome of activated T cells, providing a mechanism to balance local protein synthesis and degradation. CCT assists the folding of proteins coming fromde novosynthesis, therefore favoring mRNA translation. The functional role of this chaperonin in regulating cytoskeletal composition and dynamics at the immune synapse is discussed.

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The chaperonin CCT controls T cell receptor–driven 3D configuration of centrioles

Science Advances ◽

10.1126/sciadv.abb7242 ◽

2020 ◽

Vol 6 (49) ◽

pp. eabb7242

Author(s):

N. B. Martin-Cofreces ◽

F. J. Chichon ◽

E. Calvo ◽

D. Torralba ◽

E. Bustos-Moran ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Receptor ◽

T Cell Activation ◽

Cell Activation ◽

De Novo ◽

Lymphocyte Activation ◽

Cell Receptor ◽

Membrane Receptors ◽

Fine Tuning

T lymphocyte activation requires the formation of immune synapses (IS) with antigen-presenting cells. The dynamics of membrane receptors, signaling scaffolds, microfilaments, and microtubules at the IS determine the potency of T cell activation and subsequent immune response. Here, we show that the cytosolic chaperonin CCT (chaperonin-containing TCP1) controls the changes in reciprocal orientation of the centrioles and polarization of the tubulin dynamics induced by T cell receptor in T lymphocytes forming an IS. CCT also controls the mitochondrial ultrastructure and the metabolic status of T cells, regulating the de novo synthesis of tubulin as well as posttranslational modifications (poly-glutamylation, acetylation, Δ1 and Δ2) of αβ-tubulin heterodimers, fine-tuning tubulin dynamics. These changes ultimately determine the function and organization of the centrioles, as shown by three-dimensional reconstruction of resting and stimulated primary T cells using cryo-soft x-ray tomography. Through this mechanism, CCT governs T cell activation and polarity.

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