Actin Dynamics and Membrane Topography During T Lymphocyte Spreading

Abstract

Spreading of cells over substrates involves large scale physical rearrangements of the actin cytoskeleton and cell membrane and is of widespread physiological significance. When a T lymphocyte encounters an antigen presenting cell (APC) which presents antigen that the T cell recognizes, it spreads onto the surface of the target cell. This spreading results in signaling events and the subsequent formation of the immunological synapse. While early signaling events are known to play an important role in transcriptional response of the cell, the biophysical determinants of cell spreading kinetics are not well understood. In this study, a glass substrate coated with anti-CD3 antibodies was used to initiate the spreading response of T cells. The contact area between the cell and the substrate was used as a parameter to study the role of actin, myosin II, antigen density, membrane tension, RhoA and Rac signaling in determining the kinetics of the initial spreading response. We found that the contact area growth in time can be well described by a common physico-chemical mechanism and that the rate of spreading is independent of antibody concentration, myosin II activity and Rho signaling. We imaged the dynamics of the actin cytoskeleton using TIRF microscopy. Under certain conditions, we observed dramatic fluctuations of the edge velocity and the formation of membrane waves driven by actin polymerization at the cell substrate interface. Membrane deformations induced by such wavelike organization of the cytoskeleton may be a general phenomenon that underlies cell movement and cell-substrate interactions. Finally, we studied cell spreading on elastic substrates to investigate the possible roles of substrate rigidity on the spreading behavior, and the magnitude of traction forces exerted by the cell.