title: Two distinct functions of the carboxyl-terminal tail domain of NF-M upon neurofilament assembly: cross-bridge formation and longitudinal elongation of filaments.
abstract: Neurofilaments are the major cytoskeletal elements in the axon that take highly ordered structures composed of parallel arrays of 10-nm filaments linked to each other with frequent cross-bridges, and they are believed to maintain a highly polarized neuronal cell shape. Here we report the function of rat NF-M in this characteristic neurofilament assembly. Transfection experiments were done in an insect Sf9 cell line lacking endogenous intermediate filaments. NF-L and NF-M coassemble to form bundles of 10-nm filaments packed in a parallel manner with frequent cross-bridges resembling the neurofilament domains in the axon when expressed together in Sf9 cells. Considering the fact that the expression of either NF-L or NF-M alone in these cells results in neither formation of any ordered network of 10-nm filaments nor cross-bridge structures, NF-M plays a crucial role in this parallel filament assembly. In the case of NF-H the carboxyl-tail domain has been shown to constitute the cross-bridge structures. The similarity in molecular architecture between NF-M and NF-H suggests that the carboxyl-terminal tail domain of NF-M also constitutes cross-bridges. To examine this and to further investigate the function of the carboxyl-terminal tail domain of NF-M, we made various deletion mutants that lacked part of their tail domains, and we expressed these with NF-L. From this deletion mutant analysis, we conclude that the carboxyl-terminal tail domain of NF-M has two distinct functions. First, it is the structural component of cross-bridges, and these cross-bridges serve to control the spacing between core filaments. Second, the portion of the carboxyl-terminal tail domain of NF-M that is directly involved in cross-bridge formation affects the core filament assembly by helping them to elongate longitudinally so that they become straight.
authors.affiliation: Department of Anatomy and Cell Biology, University of Tokyo, Faculty of Medicine, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Nakagawa
authors.forename: J
authors.initials: J
authors.lastname: Chen
authors.forename: Z
authors.initials: Z
authors.lastname: Zhang
authors.forename: Y
authors.initials: Y
authors.lastname: Kanai
authors.forename: N
authors.initials: N
authors.lastname: Hirokawa
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D002460:Cell Line; D003599:Cytoskeleton / Q000648:ultrastructure; D005822:Genetic Vectors / Q000235:genetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D016900:Neurofilament Proteins / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure*; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D051381:Rats; D017384:Sequence Deletion / Q000502:physiology; D018411:Spodoptera
chemical_list: D016900:Neurofilament Proteins; D016271:Proto-Oncogene Proteins c-myc; C044882:neurofilament protein L; C061195:neurofilament protein M