The function and intracellular localisation of the non-catalytic NH2-terminal region of focal adhesion kinase (FAK) are unclear. We investigated the targetting of the FAK NH2-terminal domain in HEK 293 and epithelial MDCK cells. Exogenous expression of a variety of GFP-fused and epitope-tagged NH2 terminal domain constructs either including or lacking the major Tyr 397 autophosphorylation and Src-binding site targeted to nuclei and cell-cell junctions in HEK 293 cells and co-localised at junctions with occludin, and beta1 integrin subunits at junctions. Mutation of Tyr 397 also had no effect on localisation of the NH2-terminal domain. In contrast, constructs encoding either the kinase or focal adhesion targeting (FAT) domains but lacking the NH2-terminal region failed to localise to intercellular junctions or nuclei. The NH2-terminal domain was not associated with beta1 integrin subunits as indicated by co-immunoprecipitation experiments, but did co-localise with cortical actin filaments. The NH2-terminal domain also targetted to nuclei and intercellular junctions in MDCK cells, whereas full-length FAK localised only to focal adhesions in these cells. These results indicate that the FAK NH2-terminal domain targets to epithelial intercellular junctions and nuclei and suggest novel functions for FAK NH2-terminal domain fragments independent of Y397, kinase, and FAT domains. (C) 2002 Elsevier Science (USA). All rights reserved.