Abstract
The mammalian focal adhesion kinase (FAK) family of nonreceptor protein-tyrosine kinases have been implicated in controlling a multitude of cellular responses to the engagement of cell surface integrins and G protein-coupled receptors. We describe here a Drosophila melanogaster FAK homologue, DFak56, which maps to band 56D on the right arm of the second chromosome. Full-length DFak56 cDNA encodes a phosphoprotein of 140 kDa, which shares strong sequence similarity not only with mammalian p125(FAK) but also with the more recently described mammalian Pyk2 (also known as CAKbeta, RAFTK, FAK2, and CADTK) FAK family member. DFak56 has intrinsic tyrosine kinase activity and is phosphorylated on tyrosine in vivo. As is the case for FAK, tyrosine phosphorylation of DFak56 is increased upon plating Drosophila embryo cells on extracellular matrix proteins. In situ hybridization and immunofluorescence staining analysis showed that DFak56 is ubiquitously expressed with particularly high levels within the developing central nervous system. We utilized the UAS-GAL4 expression system to express DFak56 and analyze its function in vivo. Overexpression of DFak56 in the wing imaginal disc results in wing blistering in adults, a phenotype also observed with both position-specific integrin loss of function and position-specific integrin overexpression. Our results imply a role for DFak56 in adhesion-dependent signaling pathways in vivo during D. melanogaster development.
Highlights
The mammalian focal adhesion kinase (FAK) family of nonreceptor protein-tyrosine kinases have been implicated in controlling a multitude of cellular responses to the engagement of cell surface integrins and G proteincoupled receptors
The kinase domain of DFak56 (Fig. 1C, shaded) contains several sequence motifs conserved among protein-tyrosine kinase (PTK), including the tripeptide motif DFG that is found in most protein kinases, and a consensus ATP-binding motif GXGXXG followed by an AXK sequence downstream (Fig. 1C, underlined)
The N-terminal domain of DFak56 contains a YAEI consensus Src SH2 domain-binding sequence starting at Tyr430 (Fig. 1C, boxed), identical to the YAEI motifs in FAK and Pyk2, which bind to the SH2 domain of mammalian Src when phosphorylated
Summary
The mammalian focal adhesion kinase (FAK) family of nonreceptor protein-tyrosine kinases have been implicated in controlling a multitude of cellular responses to the engagement of cell surface integrins and G proteincoupled receptors. FAK is the founder member of a structurally conserved family of cytoplasmic nonreceptor protein-tyrosine kinases implicated in controlling cellular responses to the engagement of cell surface receptors This protein-tyrosine kinase (PTK) subfamily so far comprises two mammalian members: FAK and Pyk ( known as CAK, RAFTK, FAK2, and CADTK) [2,3,4,5,6,7,8,9]; they have 45% overall sequence identity, contain a central catalytic domain flanked by large N- and C-terminal domains, and possess a conserved Src SH2 binding autophosphorylation site (FAK ϭ Tyr397; Pyk ϭ Tyr402). Transient overexpression of the noncatalytic C-terminal domain of FAK, FRNK, inhibits FN-stimulated cell spreading, inhibits the tyrosine phosphorylation of targets such as paxillin [27, 28], and re-
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