TROPOMODULIN CAPS THE POINTED ENDS OF ACTIN-FILAMENTS

Many proteins have been shown to cap the fast growing (barbed) ends of actin filaments, but none have been shown to block elongation and dep olymerization at the slow growing (pointed) filament ends. Tropomoduli n is a tropomyosin-binding protein originally isolated from red blood cells that has been localized by immunofluorescence staining to a site at or near the pointed ends of skeletal muscle thin filaments (Fowler , V. M., M. A., Sussman, P. G. Miller, B. E. Flucher, and M. P. Daniel s. 1993. J. Cell Biol. 120: 411-420). Our experiments demonstrate that tropomodulin in conjunction with tropomyosin is a pointed end capping protein: it completely blocks both elongation and depolymerization at the pointed ends of tropomyosin-containing actin filaments in concent rations stoichiometric to the concentration of filament ends (K-d less than or equal to 1 nM). In the absence of tropomyosin, tropomodulin a cts as a ''leaky'' cap, partially inhibiting elongation and depolymeri zation at the pointed filament ends (Kd for inhibition of elongation = 0.1-0.4 mu M). Thus, tropomodulin can bind directly to actin at the p ointed filament end. Tropomodulin also doubles the critical concentrat ion at the pointed ends of pure actin filaments without affecting eith er the rate or extent of polymerization at the barbed filament ends, i ndicating that tropomodulin does not sequester actin monomers. Our exp eriments provide direct biochemical evidence that tropomodulin binds t o both the terminal tropomyosin and actin molecules at the pointed fil ament end, and is the long sought-after pointed end capping protein. W e propose that tropomodulin plays a role in maintaining the narrow len gth distributions of the stable, tropomyosin-containing actin filament s in striated muscle and in red blood cells.