A novel component of the axonal cortical cytoskeleton, A60, defined by a monoclonal antibody

A Mr 60,000 protein of the axonal cortical cytoplasm, which is recognized by a novel monoclonal antibody, is described. The antibody, DR1, was produced by immunizing mice with a soluble extract of bovine brain membranes that is enriched in known membrane cytoskeletal proteins. DR1 recognizes a Mr 60,000 protein in this extract. Immunofluorescence and subcellular fractionation reveal that the protein is primarily located in axons, where it appears to form a thick lining to the axolemma. Operationally, this Mr 60,000 protein is defined as a cytoskeleton-associated peripheral membrane protein. It is solubilized from brain membranes only under harsh conditions (0.1 M-NaOH), but not with KI (0.8 M) or Triton X-100 (1%). It is present at higher levels in the central nervous system than in peripheral nerves that have been examined. The Mr 60,000 protein copurifies with neurofilaments through cycles of assembly and disassembly. It does not appear to react with the anti-IFA antibody, suggesting that it is not a member of the intermediate filament class of proteins. This Mr 60,000 protein, which we designate A60, is distinct from other known neurofilament-associated proteins, including the Mr 60,000 protein alpha-internexin and the Mr 58,000 intermediate filament protein peripherin. A60 is suggested as being a previously unrecognized component of the axonal cortical cytoskeleton.