Macrophage caldesmon is an actin bundling protein.
A rapid purification procedure was developed for the isolation of caldesmon (CaD) from rabbit alveolar macrophage. The purified protein migrated with an apparent M(r) of 74,000 +/- 4000 on SDS-PAGE and cross-reacted with anti-gizzard CaD antibodies. A higher M(r) isoform was isolated from chicken gizzard. Their actin-binding parameters and effects on actomyosin-ATPase activity were investigated under identical experimental conditions. Electron microscope studies revealed that macrophage CaD was able to cross-link actin filaments into both networks and bundles. Compact F-actin bundles were predominantly or exclusively seen at cross-linker to actin molar ratios in the 1:20 to 1:10 range. Apparent K(a) at extrapolated saturation of the CaD-binding sites on F-actin was 1.2 x 10(6) M(-1) for macrophage CaD and 1.6 x 10(6) M(-1) for chicken gizzard CaD. CaD from either source was able to stimulate the actin-activated ATPase activity of macrophage myosin. Unexpectedly, chicken gizzard CaD also increased the ATPase activity of gizzard myosin. The degree of stimulation was approximately doubled in the presence of a large excess of Ca(2+)-calmodulin but was unaffected by the presence of macrophage tropomyosin. However, macrophage CaD did not behave as a Ca(2+)- and calmodulin-regulated actin-binding protein. These results, together with published data on other well-characterized actin bundling proteins, suggest that nonmuscle CaD could be essentially involved in the formation and organization of actin bundles at adhesion sites and cell surface projections. However, they afforded no evidence that the macrophage isoform might play a specific role in the Ca(2+)-dependent regulation of actin and myosin II interactions.
Arias
M P
MP
Centre de Recherches de Biochimie Macromoléculaire, Centre National de la Recherche Scientifique, UPR 1086, 34293, Montpellier Cedex 5, France.
Pacaud
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M
D000199:Actins / Q000737:chemistry / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000494:pharmacology; D002148:Calmodulin-Binding Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D002645:Chickens; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008264:Macrophages / Q000737:chemistry* / Q000378:metabolism; D016676:Macrophages, Alveolar / Q000737:chemistry; D008854:Microscopy, Electron; D024462:Myosin Type II / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms; D011817:Rabbits
D000199:Actins; D002148:Calmodulin-Binding Proteins; D020033:Protein Isoforms; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D024462:Myosin Type II; D002118:Calcium