HS1 and Itk cooperate to regulate actin remodeling and Ca2+ signaling in T cells (35.28)

Abstract

Abstract Productive T cell activation requires efficient reorganization of the actin cytoskeleton. We have shown that the actin regulatory protein HS1 is required for the stabilization of F-actin and Vav1 at the immunological synapse (IS) and for efficient Ca2+ responses. The Tec family kinase Itk also regulates actin responses, in part by facilitating the localization of Vav1 at the IS, suggesting that these proteins act in the same pathway. Using video microscopy, we found that T cells lacking Itk or HS1 behaved similarly, extending unstable actin-rich protrusions upon TCR stimulation. HS1 and Itk could be coimmunoprecipitated from T cell lysates, and GST-pulldown studies showed that Itk's SH2 domain binds directly to two phosphotyrosines in HS1. In the absence of Itk, HS1 failed to localize to the IS. Since Itk is required for PLCγ1 phosphorylation and Ca2+ store release, we examined the Ca2+ signaling pathway in HS1 knockout T cells in greater detail. T cells lacking HS1 exhibited abnormal Ca2+ store release. These cells showed normal phosphorylation but altered localization of PLCγ1. We conclude that Itk-dependent recruitment of HS1 is required for actin stabilization at the IS, and directs the localization of PLCγ1. Supported by NIH F31-AI071385 to EC and NIH R01-A1065644 to JKB