Figure 2.

Site-specific recognition of serine-phosphorylated residues in the C terminus of FAK by phospho-specific antibodies raised against each site of phosphorylation. (A) Myc-tagged wild-type FAK or FAK variants harboring mutations at pS2 or pS3 were transfected into HeLa cells. Lysates of transfected cells were immunoblotted with anti-myc (lanes 1-3), anti-pS2 (lanes 4-6), or anti-pS3 (lanes 7-9) to demonstrate that mutation of the immunogenic phosphorylated serine residue abolished epitope recognition by the phospho-specific antibodies. Similarly, in B, FLAG-tagged FAK or FAK variants harboring mutations at pS1 or pS4 were tested with the anti-FLAG (lanes 1-3), anti-pS1 (lanes 4-6), or anti-pS4 (lanes 7-9) antibodies. Endogenous FAK is indicated by the open arrows, ectopically expressed FAK by the closed arrows.