rs. Basal fluctuations in TCR density were observed as highly dynamic structures similar in size to microclusters, but which could not be tracked from one frame to the next (Varma et al., 2006). In order to maintain low basal activity the membrane domains in which the TCR moves prior to ligand binding will likely be highly acidic and fluid.
The lamellipodium is a relatively flat membrane region with a dense mesh of branched actin filaments translocating away from the leading edge (Svitkina et al., 1997). The mesh size of the actin gel if relatively fine, with interstitial areas on the order of 100 nm based on electron microscopy analysis of model lamellipodia (Svitkina et al., 1997). This F-actin mesh is similar in scale to the lipids domains that have been identified in membranes as lipid rafts (Sharma et al., 2004). The spatial distribution of acidic lipids like PS on the inner leaflet is not known, but structural constrains suggest that liquid ordered domains should be enriched in neutral phosphatidylethanolamine (Brown and London, 2000). I would speculate that TCR maybe confined to approximately 10% of the plasma membrane composed of disordered, fluid phase and acidic lipids, which are corralled by the mobile actin elements. In the basal state the TCR cannot be individually transported into less acidic liquid ordered domains (Fig 2). Clustering of the receptors by serial ligand binding can stabilize the interaction with the F-actin meshwork and create a sufficient force to overcome resistance to entry into less acidic domains, in which the ITAMs are exposed in the presence of Src family kinases and with depletion of CD45 together lead to triggering of the signaling cascade (Fig 2). It is likely the the “lipid rafts” also anchored to disctinct cortical actin networks and this actin may anchor these domains against movement by the dendritic actin network inivolved in the retrograde flow (Chichili and Rodgers, 2007). This model invokes shearing action of the lamellipodial actin to generate force to overcome energetic barriers, similar to recent models for the role of lateral mechanical forces in integrin activation (Zhu et al., 2008).
The specific adapters that link TCR to F-actin are not well described. The complex may include NCK, SLP-76, ADAP, SKAP55, EVL, Vav, Rac, Cdc42 and WASP certainly may play a role once signaling is initiated (Barda-Saad et al., 2005; Bubeck Wardenburg et al., 1998; Krause et al., 2000). It has also been suggested that ZAP-70 may play a role through binding of ezrin (Ilani et al., 2007). It appears that TCR signaling in microclusters ceases when the microcluster reaches the actin depleted cSMAC. Since activated TCR appear equipped to generate their own dynamic actin focus, the suppression of this mode of actin dymanics in the cSMAC is likely to be active, rather than a passive dissipation of lamellar actin. Less is known about the basal interactions of TCR with actin that might be available to stabilize a cluster early in the triggering process. This is likely a complex situation. An example of the potential complexity can be seen in interaction networks recently compiled as the integrin adhesome (Zaidel-Bar et al., 2007).
Once the ITAMs are exposed Src family kinases can then initiate phosphorylation by diffusion into the same domain (Douglass and Vale, 2005)(Fig 2). Exclusion of CD45 in the nascent foci would increase the half-life of the phosphorylated ITAM allowing sufficient time for ZAP-70 recruitment (Varma et al., 2006). This cluster may interact more strongly with LAT so that ZAP-70 could phosphorylate LAT and Itk to initiate PLC-g recruitment and initiation of Ca2+ signaling, PKC activation and Ras-GRP recruitment. Integrin microclusters will be interspersed with the TCR microclusters. Integrin dependent signaling will generate additional diacylglycerol for RasGRP by activating phospholipase D and phosphatidic acid phosphatase (Mor et al., 2007). Electron micrographs of membrane sheets suggest that immunoreceptor rich and LAT rich islands only partially mix (Lillemeier et al., 2006;