Modulation of gelsolin-induced actin-filament severing by caldesmon and tropomyosin and the effect of these proteins on the actin activation of myosin Mg(2+)-ATPase activity.

We have investigated the cumulative effects of three smooth-muscle actin-binding proteins, gelsolin, caldesmon and tropomyosin, on actin activation of myosin Mg(2+)-ATPase activity under low-ionic-strength conditions. A combination of tropomyosin (at a stoicheiometric ratio to actin) and gelsolin (at a molar ratio to actin of up to 1:100) showed essentially additive stimulatory effects that were counteracted by caldesmon. Suppression of the gelsolin-induced activation of the ATPase by caldesmon was higher in the presence of tropomyosin although it was not complete even at stoicheiometric amounts of both proteins to actin.

Complementary distributions of vinculin and dystrophin define two distinct sarcolemma domains in smooth muscle.

The sarcolemma of the smooth muscle cell displays two alternating structural domains in the electron microscope: densely-staining plaques that correspond to the adherens junctions and intervening uncoated regions which are rich in membrane invaginations, or caveolae. The adherens junctions serve as membrane anchorage sites for the actin cytoskeleton and are typically marked by antibodies to vinculin. We show here by immunofluorescence and immunoelectron microscopy that dystrophin is specifically localized in the caveolae-rich domains of the smooth muscle sarcolemma, together with the caveolae-associated molecule caveolin.

Caldesmon-induced polymerization of actin from profilactin.

We have investigated the effect of caldesmon, a Ca2+/calmodulin-regulated actin-binding protein, on the complex between profilin and G-actin (profilactin). We found that smooth muscle caldesmon dissociates this complex rapidly and induces the polymerization of the released actin. Native profilactin (e.

The effect of caldesmon on dynamic properties of F-actin alone and bound to heavy meromyosin and/or tropomyosin.

The effect of caldesmon on the rotational dynamics of actin filaments alone or conjugated with heavy meromyosin and/or tropomyosin has been measured by the electron paramagnetic resonance (EPR) technique using a maleimide spin label rigidly bound to Cys374 of actin. The rotation of actin protomers in filaments and the angular distribution of spin probes on actin were determined by conventional EPR spectroscopy, while torsional motions within actin filaments were detected by saturation transfer EPR measurements. Binding of caldesmon to F-actin resulted in the reduction of torsional mobility of actin filaments.

The effect of caldesmon on assembly and dynamic properties of actin.

Our earlier fluorescence measurements using N-(1-pyrenyl)iodoacetamide-labeled actin revealed that caldesmon interacts with G-actin accelerating its nucleation at low salt concentration and causing polymerization in the absence of sale [Gałazkiewicz, B., Mossakowska, M., Osińska, H.