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

We examined the localization and role of alpha-IN vs. other neuronal intermediate filaments before and during differentiation. Vimentin but not alpha-IN localized within filopodia-like neurites of undifferentiated cells. During differentiation, alpha-IN immunoreactivity accumulated within axonal neurites following vimentin but, as previously describe in neurons in situ, before the appearance of NF-L. We therefore manipulated alpha-IN synthesis, accumulation, and function in attempts to determine whether or not this intermediate filament species played a role in axonal development. Intracellular delivery of anti-alpha-IN antisense oligonucleotides and antibodies was permissive for neuritogenesis, yet compromised neurite elongation; this effect was further reflected in diminished levels of stabilized axonal microtubules. These data suggest that alpha-IN plays a role in the development of neuronal 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. Protease 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 co-precipitated from cells with alpha-IN and NF-L. Virtually all newly synthesized NF-H co-precipitated with NF-L, while only a small percentage co-precipitated with alpha-IN. Finally, NF-H or NF-M were absent from the axon hillock or perikaryal area at the base of neurites, where alpha-IN immunoreactivity is prominent. These data extend earlier cell-free demonstrations that NF-H preferentially associates with NF-L rather than alpha-IN.