NO synthase II in mouse skeletal muscle is associated with caveolin 3.
The inducible-type NO synthase (NOS II; iNOS) is constitutively expressed in slow-twitch skeletal muscle fibres of guinea-pigs [Gath, Closs, Gödtel-Armbrust, Schmitt, Nakane, Wessler and Förstermann (1996) FASEB J. 10, 1614-1620]. Here we studied the expression of NOS II in skeletal muscle of wild-type and NOS II-deficient mice and investigated the molecular basis for the membrane association of this NOS in muscle. A basal expression of NOS II mRNA and protein was detected in skeletal muscle from untreated wild-type mice; expression increased when mice were treated with bacterial lipopolysaccharide (LPS). No NOS II was found in any tissue of untreated or LPS-treated NOS II-deficient mice. Immunoprecipitation experiments were performed with homogenates of gastrocnemius muscle from untreated or LPS-treated wild-type mice. A NOS II-specific antibody precipitated caveolin 3 in all homogenates investigated, the effect being most pronounced in skeletal muscle from LPS-treated animals. Conversely, an antibody against caveolin 3 co-precipitated NOS II in muscle homogenates. Similarly, a weak co-precipitation of NOS II and caveolin 3 was seen in homogenates of untreated murine C2C12 myotubes; co-precipitation was markedly enhanced in cells stimulated with LPS/interferon gamma. The association of NOS II with caveolin 3 might have implications for the regulation of contraction of, and/or glucose uptake by, slow-twitch muscle fibres.
Gath
I
I
Department of Pharmacology, Johannes Gutenberg University, Obere Zahlbacher Strasse 67, D-55101 Mainz, Germany.
Ebert
J
J
Gödtel-Armbrust
U
U
Ross
R
R
Reske-Kunz
A B
AB
Förstermann
U
U
D000818:Animals; D015153:Blotting, Western; D051244:Caveolin 3; D022461:Caveolins; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D002531:Cerebellum / Q000187:drug effects / Q000201:enzymology; D004790:Enzyme Induction / Q000187:drug effects; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D019001:Nitric Oxide Synthase / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics / Q000378:metabolism
C495548:Cav3 protein, mouse; D051244:Caveolin 3; D022461:Caveolins; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D012333:RNA, Messenger; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase