A Proteomic Study of Myosin II Motor Proteins during Tumor Cell Migration

• Published in: Journal of molecular biology Vol. 407; no. 5; pp. 673 - 686
• Main Authors: Betapudi, Venkaiah, Gokulrangan, Giridharan, Chance, Mark R., Egelhoff, Thomas T.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 15-04-2011
• Subjects: Animals; breast neoplasms; Breast Neoplasms - metabolism; Breast Neoplasms - pathology; Casein Kinase II - chemistry; Casein Kinase II - genetics; Casein Kinase II - metabolism; Cell Line, Tumor; cell migration; cell movement; Cell Movement - physiology; Cercopithecus aethiops; COS Cells; cytoskeleton; Female; fibronectins; Fibronectins - metabolism; HeLa Cells; Humans; integrins; Models, Molecular; myosin heavy chains; Myosin Heavy Chains - chemistry; Myosin Heavy Chains - genetics; Myosin Heavy Chains - metabolism; myosin II; Myosin Type II - chemistry; Myosin Type II - genetics; Myosin Type II - metabolism; Phosphorylation; Protein Isoforms - chemistry; Protein Isoforms - genetics; Protein Isoforms - metabolism; protein kinase C; protein phosphorylation; Proteome - analysis; proteomics; Pseudopodia - metabolism; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; RNA; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; signal transduction; PBS; EGF; cytoskeleton; RLC; MHC; PKC; siRNA; CIP; GFP; LC-MS/MS; CK-II; myosin II; cell migration; phosphorylation; EIC; ERK
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2011.02.010

Myosin II motor proteins play important roles in cell migration. Although myosin II filament assembly plays a key role in the stabilization of focal contacts at the leading edge of migrating cells, the mechanisms and signaling pathways regulating the localized assembly of lamellipodial myosin II filaments are poorly understood. We performed a proteomic analysis of myosin heavy chain (MHC) phosphorylation sites in MDA-MB 231 breast cancer cells to identify MHC phosphorylation sites that are activated during integrin engagement and lamellar extension on fibronectin. Fibronectin-activated MHC phosphorylation was identified on novel and previously recognized consensus sites for phosphorylation by protein kinase C and casein kinase II (CK-II). S1943, a CK-II consensus site, was highly phosphorylated in response to matrix engagement, and phosphoantibody staining revealed phosphorylation on myosin II assembled into leading-edge lamellae. Surprisingly, neither pharmacological reduction nor small inhibitory RNA reduction in CK-II activity reduced this stimulated S1943 phosphorylation. Our data demonstrate that S1943 phosphorylation is upregulated during lamellar protrusion, and that CK-II does not appear to be the kinase responsible for this matrix-induced phosphorylation event.