The Actin Cytoskeleton Dynamically Associates with T-Cell Receptor Clusters

Abstract

The actin cytoskeleton is a key determinant of cell surface protein organization in many eukaryotic cells. In mammalian T-cells undergoing antigen-mediated activation, the cytoskeleton drives the development of a set of macroscale protein domains, collectively termed the supramolecular activation cluster. These protein domains, composed of T-cell receptors surrounded by adhesion molecules and their ligands, are highly characteristic of T-cell activation and are thought to play an important role in modulating receptor signaling intensity. Though significant research has been undertaken to elucidate the interactions among various receptors involved in T-cell activation, the nature of the interactions between these receptors and actin remains poorly established. We have used live-cell fluorescence microscopy to image the actin cytoskeleton as it interacts with T-cell receptors in real time. Our results support recent work from our lab that had shown that T-cell receptors are likely to be friction-coupled to the cytoskeleton. Actin density tracking has also extended that work by demonstrating that cytoskeletal velocity may be affected by mobility-limited T-cell receptor clusters, and thus that T-cell receptors may have the capacity to regulate actin flow. In addition, we have evaluated the time dynamics of the T-cell receptor-actin interaction and found that actin periodically accumulates and dissipates at T-cell receptor clusters. By applying an autocorrelation function to our image stacks, we found that the half-decay time (Tau1/2) of the actin fluorescence at regions corresponding to T-cell receptor clusters was significantly increased compared to background, indicating a greater persistence of actin in those regions. Thus we have developed a novel method of analyzing actin kinetics and shown that the actin cytoskeleton dynamically associates with T-cell receptor clusters.