CD28 engagement promotes actin polymerization through the activation of the small Rho GTPase Cdc42 in human T cells

Salazar-Fontana LI, Barr V, Samelson LE and Bierer BE
Source: J Immunol
Publication Date: (2003)
Issue: 171: 2225-2232
Research Area:
Immunotherapy / Hematology
Cells used in publication:
T cell, human peripheral blood unstim.
Species: human
Tissue Origin: blood
It is now generally accepted that CD28 is an important costimulator for the TCR/CD3 complex promoting a sustained signaling cascade that will allow the T cell to proliferate and function. Using Ab-coated surfaces the authors show, that CD28 engagement induce the formation of cytoplasmic elongations enriched in filamentous actin (microspikes). Small Rho GTPases, namely Cdc42, Rac, and Rho, have been associated with remodelling of the actin cytoskeleton in many cell types, including T lymphocytes. To further reveal the role of Cdc42, primary human T cells were nucleofected with expression vectors encoding a GFP fusion protein of either a dominant-negative (DN) version, or a constitutively active (CA) version of Cdc42. Cells were plated on coverslips coated with CD28 or PLL and stained for F-actin. Immunofluorescence images show that Cdc42 is responsible of CD28-induced actin reorganization and the formation of microspikes.
Engagement of the costimulatory molecule CD28 is an important step in the optimal activation of T cells. Nevertheless, the specific role of CD28 in the formation of the immunological synapse and cytoskeletal changes that occur upon TCR/CD3 complex engagement is still poorly understood. Using Ab-coated surfaces, we show that CD28 engagement in the absence of any other signal induced the formation of cytoplasmic elongations enriched in filamentous actin (F-actin), in this work called filopodia or microspikes. Such structures were specific for engagement of CD28 on mAb-coated surfaces because they could not be observed in surfaces coated with either poly(L-lysine) or anti-CD3 mAb. The signaling pathway coupling CD28 to cytoskeletal rearrangements required Src-related kinase activity and promoted Vav phosphorylation and Cdc42 activation independently of the zeta-chain-associated kinase (ZAP-70). CD28-induced filopodia required Cdc42 GTPase activity, but not the related Rho GTPase Rac1. Moreover, Cdc42 colocalized to areas of increased F-actin. Our results support a specific role for the activation of the small Rho GTPase Cdc42 in the actin reorganization mediated by CD28 in human T cells.