Interactions of short mimetic peptides from the intermediate filament protein desmin with filaments in vitro and in vivo

Kurzfassung

The intermediate filament protein desmin is an important protein for the maintenance of structure and functionality in muscle cells. Mutations lead to myofibrillar myopathies in humans and in animal models, with both skeletal and cardiac muscle being affected. Desmin displays a tripartite structure consisting of a central a-helical domain, framed by two intrinsically disordered terminal domains. The functions of both terminal domains are still not fully understood. In this study, the interactions of these amino- and carboxy-terminal unstructured domains within the filament in vitro as well as in transfected cells were evaluated by using five different mimetic peptides. The peptides were selected from evolutionarily highly conserved sequence elements in the tail and in the head domain, respectively. We hypothesized that these peptide motifs may therefore be important for intra- and inter-filament interactions, and thus are taking part in the maintenance of the mechanical properties of intermediate filament networks. To investigate their potential interactions with desmin in vitro, we co- assembled fluorescently labeled peptides with recombinant desmin filaments. Accordingly, we studied the interactions in vivo by transfecting the myoblasts cells line C2C12 with the cDNA of the peptides coupled to fluorescent proteins. With these approaches, we wanted to achieve a better understanding of these domains regarding their role in filament formation, the regulation of the turnover of mature filaments as well as their interaction with cellular organelles in the living cell. Our findings revealed that the C- terminal peptides showed low binding affinity to desmin filaments both in vitro and in vivo. In contrast, our studies with the N-terminal peptide revealed that this domain is an essential element for assembly initiation and filament elongation, as previously described for vimentin and keratin. As for the tail peptides, no pronounced interaction of the head peptide with mature filaments was observed in C2C12 cells after transfection.