RRC ID 68510
Author Ke X, Terashima M, Nariai Y, Nakashima Y, Nabika T, Tanigawa Y.
Title Nitric oxide regulates actin reorganization through cGMP and Ca(2+)/calmodulin in RAW 264.7 cells.
Journal Biochim Biophys Acta
Abstract Nitric oxide (NO) has been reported to be involved in the regulation of pseudopodia formation, phagocytosis and adhesion in macrophages through the reorganization of actin. In the present study, we directly separated the globular (G) and filamentous (F) actin from quiescent or NO-stimulated macrophage-like cell line RAW 264.7 cells in order to investigate the dynamic redistribution of actin pools. We also focused on the regulatory mechanisms of actin assembly, induced by NO and its possible subsequent signaling pathway. We showed that predominant G-actin coexisted with Triton X-100-insoluble filamentous (TIF) and Triton X-100-soluble filamentous actin in resting RAW 264.7 cells. The exogenous NO produced by (+/-)-(E)-2-[(E)-hydroxyimino]-6-methoxy-4-methyl-5-nitro-3-hexenamide (NOR1), the endogenous NO induced by lipopolysaccharide (LPS) plus interferon-gamma (IFNgamma), and dibutyryl-cGMP increased the contents of TIF-actin in dose- and time-dependent manners and altered its morphology. The increase in the TIF-actin contents induced by NOR1 or LPS plus IFNgamma was efficiently blocked by the radical scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide and the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one or the arginine analogue N(G)-monomethyl-L-arginine acetate, respectively. Preincubation with the calmodulin antagonist W-7 almost completely blocked the NO-induced TIF-actin increase and morphological change. On the other hand, preincubation with C3 transferase, an inhibitor of Rho protein, efficiently prevented the change in cell morphology, but had no effect on the TIF-actin increase. We postulate that cGMP and subsequent Ca(2+)/calmodulin may be key regulators of actin reorganization in NO-stimulated RAW 264.7 cells.
Volume 1539(1-2)
Pages 101-13
Published 2001-5-28
DOI 10.1016/s0167-4889(01)00090-8
PII S0167-4889(01)00090-8
PMID 11389972
MeSH Actins / analysis Actins / metabolism* Animals Bucladesine / pharmacology Calcium / metabolism* Calmodulin / antagonists & inhibitors Calmodulin / metabolism* Cell Line Cyclic GMP / analysis Cyclic GMP / metabolism* Dibutyryl Cyclic GMP / pharmacology Enzyme Inhibitors / pharmacology Flow Cytometry Interferon-gamma / pharmacology Lipopolysaccharides / pharmacology Mice Nitric Oxide / pharmacology* Nitric Oxide Donors / pharmacology Oxadiazoles / pharmacology Quinoxalines / pharmacology Sulfonamides / pharmacology
IF 3.411
Human and Animal Cells RAW 264(RCB0535)