Photoinduced inactivation of protein kinase C by dequalinium identifies the RACK-1-binding domain as a recognition site

1,1'-Decamethylenebis-4-aminoquinaldinium diiodide (DECA; dequalinium) is an anti-tumor agent and protein kinase C (PKC) inhibitor whose mechanism of action with PKC is unknown, This study reports that with human PKC alpha, DECA exhibited competitive inhibition (K-i = 11.5 +/- 5 mu M) with respect to RACK-1 (receptor for activated C kinase-1), an adaptor protein that has been proposed to bind activated PKC following translocation (Ron, D., Luo, J., and Mochly-Rosen, D. (1995) J. Biol. Chem. 270, 24180-24187), When exposed to UV light, DECA covalently modified and irreversibly inhibited PKC (alpha or beta), with IC50 = 7-18 mu M. UV/DECA treatment of synthetic peptides modeled after the RACK-1-binding site in the C2 region of PKC beta induced modification of Ser(218)-Leu-Asn-Pro-Glu-Trp-Asn-Glu-Thr(226), but not of a control peptide, This modification occurred ata tryptophan residue (Trp(223)) that is conserved in all conventional PKC isoforms. In overlay assays with native RACK-1 that had been immobilized on nitrocellulose, UV-treated control PKC alpha bound well to RACK-1, whereas UV/DECA-inactivated PKC alpha had reduced binding activity, The significance of these findings is shown with adenocarcinoma cells, which, when pretreated with 10 mu M DECA and UV light, exhibited diminished 12-O-tetradecanoylphorbol-13-acetate-induced PKC alpha translocation. Overall, this work identifies DECA as a tool that prevents PKC translocation by inhibiting formation of the PRC RACK-1 complex.