Thrombin-induced phosphorylation of MARCKS does not alter its interactions with calmodulin or actin

• Published in: Cellular Signalling 2000-02, Vol.12 (2), p.71-79
• Main Authors: Neltner, Bonnie S., Zhao, Ying, Sacks, David B., Davis, Harold W.
• Format: Article
• Language: English
• Subjects: Actin; Actins - analysis; Actins - metabolism; Animals; Calmodulin; Calmodulin - analysis; Calmodulin - metabolism; Cattle; CHO Cells; Cricetinae; Cytoskeleton - chemistry; Cytoskeleton - drug effects; Cytoskeleton - metabolism; DNA, Complementary; Endothelial cells; Endothelium, Vascular - cytology; Endothelium, Vascular - drug effects; Endothelium, Vascular - metabolism; Hemostatics - pharmacology; Intracellular Signaling Peptides and Proteins; MARCKS; MARCKS protein; Membrane Proteins; Myosin light chains; Myosin Light Chains - metabolism; Myristoylated Alanine-Rich C Kinase Substrate; Phosphorylation; Proteins - analysis; Proteins - genetics; Proteins - metabolism; Pulmonary Artery - cytology; Signal Transduction - drug effects; Signal Transduction - physiology; Thrombin - pharmacology; Transfection
• ISSN: 0898-6568; 1873-3913
• DOI: 10.1016/S0898-6568(99)00065-0

Myristoylated alanine-rich C kinase substrate (MARCKS) is a calmodulin (CaM)- and actin-binding protein and prominent protein kinase C (PKC) substrate. In vitro phosphorylation of MARCKS by PKC has been shown to induce the release of both CaM and actin, leading to the suggestion that MARCKS may regulate CaM availability during agonist-induced signalling. In support of this hypothesis we previously demonstrated that thrombin-induced MARCKS phosphorylation in endothelial cells (EC) parallels activation of myosin light chain kinase, a CaM-dependent enzyme. To test this theory further, we transfected CHO cells, which normally do not express significant levels of MARCKS, with a MARCKS cDNA. The thrombin-stimulated phosphorylation of myosin light chains and the sensitivity to CaM antagonists in the MARCKS overexpressing cells was the same as that in control CHO cells. MARCKS associated with the actin cytoskeleton in EC was markedly increased upon treatment with the PKC activator, PMA, but only modestly enhanced by thrombin treatment. Similarly, colocalisation of MARCKS with actin was enhanced when the EC were challenged with PMA but not thrombin. These data may be partially explained by PKC-independent phosphorylation of MARCKS in response to thrombin stimulation.