Dynamic microtubules regulate cellular contractility during T-cell activation

Abstract

T-cell receptor (TCR) triggering and subsequent T-cell activation are essential for the adaptive immune response. Recently, multiple lines of evidence have shown that force transduction across the TCR complex is involved during TCR triggering, and that the T cell might use its force-generation machinery to probe the mechanical properties of the opposing antigen-presenting cell, giving rise to different signaling and physiological responses. Mechanistically, actin polymerization and turnover have been shown to be essential for force generation by T cells, but how these actin dynamics are regulated spatiotemporally remains poorly understood. Here, we report that traction forces generated by T cells are regulated by dynamic microtubules (MTs) at the interface. These MTs suppress Rho activation, nonmuscle myosin II bipolar filament assembly, and actin retrograde flow at the T-cell-substrate interface. Our results suggest a novel role of the MT cytoskeleton in regulating force generation during T-cell activation.T lymphocytes, central players in the adaptive immune response, are activated when T-cell receptors (TCRs) on their surface recognize cognate peptide-major histocompatibility complex (pMHC) expressed on the surface of antigen-presenting cells (APCs). A burst of actin polymerization is triggered upon TCR stimulation (1), leading to an enhancement of the cell/APC contact area as the T cell spreads over the surface of the APC (2) and the formation of a macromolecular protein assembly known as the immunological synapse (IS) (3, 4). Accompanying IS formation, T cells also undergo a rapid polarization of the microtubule (MT) cytoskeleton, within 1-2 min after initial contact, that facilitates directional secretion of cytokines and cytolytic factors toward the APC (5-7). Therefore, the contact zone between T cells and APCs is a site at which the MT and actin cytoskeletons could potentially interact to regulate signaling.Recent studies have established that T cells generate significant traction stresses at the cell-cell interface, albeit relatively weak compared with adherent cells (8-11). These forces, which peak 5-10 min after stimulation, facilitate T-cell activation, in part, by inducing conformational changes in the TCR-CD3 complex (12-15). Although actin polymerization/depolymerization dynamics are essential for T cells to maintain dynamic traction stresses and to drive calcium influx and integrin affinity maturation (9, 16, 17), the regulatory pathways that control these cytoskeletal forces are not completely understood. In particular, whether and how the polarized MT cytoskeleton interacts with the actin cytoskeleton and regulates force generation at the T-cell-APC contact remain open questions.MTs in the cell exist in two populations: dynamic/tyrosinated MTs and stable MTs that have undergone posttranslational modifications, including detyrosination and acetylation (18). Previous studies of T-cell activation have elucidated that microtubuleorganizing center (MTOC) translocation is associated with the formati...

Ex vivo induction of regulatory T cells from conventional CD4+ T cells is sensitive to substrate rigidity

“…A similar result was reported recently with a mesh format . These works were followed by many studies on lymphocyte responses to varying substrate rigidities which examine B cell activation, actin dynamics, and microtubule organizations of T cells …”

Enhanced Activation and Expansion of T Cells Using Mechanically Soft Elastomer Fibers

“…[14] Complementary studies showed that T cells can apply significant traction forces to an underlying substrate through the TCR complex. [15] This suggests a feedback loop through which resistance to cell-generated forces sustains activation, allowing for rigidity sensing; in this direction, actomyosin contractility is required for a range of T cell functions including mechanosensing and overall stability of cellular interfaces. [16] Identification of the specific molecular linkages between these stages is an area of active research.…”

Mechanosensing in the immune response

“…However, the contribution of the MT to force generation at the synapse has been relatively unknown, despite the fact that organelle movement as well as MT deformation clearly indicates the existence of forces. We recently found that MT tip dynamics at the lamellipodial/lamellar region modulate NMII filament assembly and lamellipodial actin flow through the RhoGTPase pathway and modulates the traction forces exerted by T cells [60]. These studies suggest that centrosomal movement and microtubule deformations result from forces generated by MT motors and the actomyosin cytoskeleton [61–64], and point to the importance of interactions between the actin and MT cytoskeleton [65] in force generation and force balance at the immune synapse, but its role in TCR signaling remains to be explored.…”

“…These forces were specific to TCR-ligand binding, and both actin dynamics (polymerization and depolymerization) and myosin activity were required for force generation. We have recently shown that microtubule tip dynamics modulate force generation in T cells [60]. …”