Neuronal intermediate filament protein alpha-internexin facilitates axonalneurite elongation in neuroblastoma cells

We examined the localization and role of alpha-IN vs. other neuronal interm ediate filaments before and during differentiation. Vimentin but not alpha- IN localized within filopodia-like neurites of undifferentiated cells. Duri ng differentiation, alpha-IN immunoreactivity accumulated within axonal neu rites following vimentin but, as previously describe in neurons in situ, be fore the appearance of NF-L. We therefore manipulated alpha-IN synthesis, a ccumulation, and function in attempts to determine whether or not this inte rmediate filament species played a role in axonal development. Intracellula r delivery of anti-alpha-IN antisense oligonucleotides and antibodies was p ermissive for neuritogenesis, yet compromised neurite elongation; this effe ct was further reflected in diminished levels of stabilized axonal microtub ules. These data suggest that alpha-IN plays a role in the development of n euronal polarity. Relatively more alpha-IN than NF-L accumulated within the plastic axonal neurites induced following serum-deprivation. while stable, dbcAMP-induced neurites treatment contained equivalent levels of each. Pro tease inhibition increased NF-L and NF-H but not alpha-IN immunoreactivity within serum-deprived neurites, suggesting that proteolysis restricts NF-L accumulation pending neurite stabilization. To test the possibility that NF -H accumulation is dependent upon NF-L and cannot be mediated by alpha-IN, we examined levels of NF-H coprecipitated from cells with alpha-IN and NF-L . Virtually all newly synthesized NF-H co-precipitated with NF-L, while onl y a small percentage co-precipitated with alpha-IN. Finally, NF-H or NF-M w ere absent from the axon hillock or perikaryal area at the base of neurites , where alpha-IN immunoreactivity is prominent. These data extend earlier c ell-free demonstrations that NF-H preferentially associates with NF-L rathe r than alpha-IN. Cell Motil. Cytoskeleton 43:322-333, 1999. (C) 1999 Wiley- Liss, Inc.