Regulation of chemotaxis by the cytoplasmic domain of tissue factor.

We previously demonstrated that FVIIa bound to tissue factor (TF) induces a hyperchemotactic response towards PDGF-BB. The aim of the present study was to investigate the role of the cytoplasmic domain of TF in cell migration. Porcine aortic endothelial (PAE) cells expressing human PDGF beta-receptors (PAE/PDGFRbeta) were transfected for expression of human wildtype TF (PAE/PDGFRbeta,TF), a construct lacking the cytoplasmic domain (PAE/PDGFRbeta,TFDeltacyto), a construct with alanine replacement of serine 258 (PAE/PDGFRbeta,TFS258A), or a construct with alanine replacement of serine 253, 258 and 263 in the cytoplasmic domain (PAE/PDGFRbeta,TF3SA). All stably transfected cell lines expressed functional TF. Chemotaxis was analyzed in a modified Boyden chamber assay. PAE/PDGFRbeta,TF cells stimulated with FVIIa migrated towards a 100-fold lower concentration of PDGF-BB than in the absence of FVIIa, however, hyperchemotaxis was not seen in PAE/PDGFRbeta,TFDeltacyto cells. PAE/PDGFRbeta/TFS258A and PAE/PDGFRbeta,TF3SA cells responded to low levels of PDGF-BB, but migrated a significantly shorter distance than PAE/PDGFRbeta,TF cells. Thus, hyperchemotaxis towards PDGF-BB is likely to depend in part on phosphorylation of the cytoplasmic domain of TF. We conclude that the cytoplasmic domain of TF plays a pivotal role in modulating cellular migration response. Our results suggest that the FVIIa/TF complex mediates intracellular signaling by alternative signal transduction pathway(s).