INHIBITION OF CAPZ DURING MYOFIBRILLOGENESIS ALTERS ASSEMBLY OF ACTIN-FILAMENTS

The actin filaments of myofibrils are highly organized; they are of a uniform length and polarity and are situated in the sarcomere in an al igned array. We hypothesized that the barbed-end actin-binding protein , CapZ, directs the process of actin filament assembly during myofibri llogenesis. We tested this hypothesis by inhibiting the actin-binding activity of CapZ in developing myotubes in culture using two different methods. First, injection of a monoclonal antibody that prevents the interaction of CapZ and actin disrupts the non-striated bundles of act in filaments formed during the early stages of myofibril formation in skeletal myotubes in culture. The antibody, when injected at concentra tions lower than that required for disrupting the actin filaments, bin ds at nascent Z-disks. Since the interaction of CapZ and the monoclona l antibody are mutually exclusive, this result indicates that CapZ bin ds nascent Z-disks independent of an interaction with actin filaments. In a second approach, expression in myotubes of a mutant form of CapZ that does not bind actin results in a delay in the appearance of acti n in a striated pattern in myofibrils. The organization of cr-actinin at Z-disks also is delayed, but the organization of titin and myosin i n sarcomeres is not significantly altered. We conclude that the intera ction of CapZ and actin is important for the organization of actin fil aments of the sarcomere.