Calcium/calmodulin-dependent protein kinase II binds to Raf-1 and modulates integrin-stimulated ERK activation.
Integrin activation generates different signalings in a cell type-dependent manner and stimulates cell proliferation through the Ras/Raf-1/Mek/Erk pathway. In this study, we demonstrate that integrin stimulation by fibronectin (FN), besides activating the Ras/Erk pathway, generates an auxiliary calcium signal that activates calmodulin and the Ca2+/calmodulin-dependent protein kinase II (CaMKII). This signal regulates Raf-1 activation by Ras and modulates the FN-stimulated extracellular signal-regulated kinase (Erk-1/2). The binding of soluble FN to integrins induced increase of intracellular calcium concentration associated with phosphorylation and activation of CaMKII. In two different cell lines, inhibition of CaMKII activity by specific inhibitors inhibited Erk-1/2 phosphorylation. Whereas CaMK inhibition affected neither integrin-stimulated Akt phosphorylation nor p21Ras or Mek-1 activity, it was necessary for Raf-1 activity. FN-induced Raf-1 activity was abrogated by the CaMKII specific inhibitory peptide ant-CaNtide. Integrin activation by FN induced the formation of a Raf-1/CaMKII complex, abrogated by inhibition of CaMKII. Active CaMKII phosphorylated Raf-1 in vitro. This is the first demonstration that CaMKII interplays with Raf-1 and regulates Erk activation induced by Ras-stimulated Raf-1. These findings also provide evidence supporting the possible existence of cross-talk between other intracellular pathways involving CaMKII and Raf-1.
Illario
Maddalena
M
Dipartimento di Biologia e Patologia Cellulare e Molecolare, Università Federico II, Napoli, 80131 Italy.
Cavallo
Anna Lina
AL
Bayer
K Ulrich
KU
Di Matola
Tiziana
T
Fenzi
Gianfranco
G
Rossi
Guido
G
Vitale
Mario
M
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D016023:Integrins / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction; D013997:Time Factors; D018631:ras Proteins / Q000378:metabolism
D004791:Enzyme Inhibitors; D005353:Fibronectins; D016023:Integrins; D019908:Proto-Oncogene Proteins c-raf; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins; D002118:Calcium