The F-actin cross-linking and focal adhesion protein filamin A is a ligand and in vivo substrate for protein kinase C alpha.
Filamin A is an established structural component of cell-matrix adhesion sites. In addition, it serves as a scaffold for the subcellular targeting of different signaling molecules. Protein kinase C (PKC) has been found associated with filamin; however, details about this interaction and its significance for cell-matrix adhesion-dependent signaling have remained elusive. We performed a yeast two-hybrid analysis using protein kinase Calpha as a bait and identified filamin as a direct binding partner. The interaction was confirmed in transfected HeLa cells, and serial truncation fragments of filamin A were employed to identify two binding sites on filamin. In vitro ligand binding assays revealed a Ca2+ and phospholipid-dependent association of the regulatory domain of protein kinase C with these sites. Phosphorylation of filamin was found to be isoform-restricted, leading to phosphate incorporation in the C termini of filamin A and C, but not B. PKC-dependent phosphorylation of filamin was also detected in cells. Our data suggest an intimate interaction between filamin and PKC in cell signaling.
Tigges
Ulrich
U
Department of Cell Biology, Zoological Institute, Technical University of Braunschweig, Braunschweig, Germany.
Koch
Bettina
B
Wissing
Josef
J
Jockusch
Brigitte M
BM
Ziegler
Wolfgang H
WH
D000199:Actins / Q000378:metabolism*; D001665:Binding Sites; D002118:Calcium; D003285:Contractile Proteins / Q000378:metabolism*; D064448:Filamins; D022001:Focal Adhesions / Q000737:chemistry; D006801:Humans; D008024:Ligands; D008840:Microfilament Proteins / Q000378:metabolism*; D010743:Phospholipids; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques
D000199:Actins; D003285:Contractile Proteins; D064448:Filamins; D008024:Ligands; D008840:Microfilament Proteins; D010743:Phospholipids; D020033:Protein Isoforms; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D002118:Calcium