Syk tyrosine kinase is required for immunoreceptor tyrosine activation motif-dependent actin assembly

Clustering of several multisubunit receptors on hematopoetic cells results in a signaling cascade involving the phosphorylation of immunoreceptor tyrosine activation motifs, or 'ITAMs,' and actin polymerization. Recent experiments indicate that direct clustering of the ITAM-binding protein, p72(syk) (Syk), is capable of transmitting a phagocytic signal in COS cells (Greenberg, S., Chang, P., Wang, D., Xavier, R., and Seed, B. (1996) Proc. Natl. Acad. Sci. U.S.A. 93, 1103-1107). However, the possibility of redundant signaling pathways makes it difficult to test the requirement for Syk in ITAM-dependent actin polymerization in hematopoetic cells. We developed a model system to study ITAM-dependent actin assembly. DT40 lymphocytes were transfected with fusion proteins encoding the transmembrane and cytosolic domains of the ITAM-containing γ subunit of Fc receptors. Clustering the γ-containing fusion proteins with IgG-coated erythrocytes triggered submembranous actin assembly. This response depended on an intact ITAM, was absent in cell lines that had been engineered to lack Syk, and was augmented in cell lines that stably overexpressed Syk. These experiments demonstrate an absolute requirement for Syk tyrosine kinase in ITAM-dependent actin assembly in transfected lymphocytes.