Microtubule-associated protein tau is required for axonal neurite elaboration by neuroblastoma cells

NB2a/d1 neuroblastoma cells constitutively express multiple isoforms of the microtubule-associated protein tau and incorporate this protein into the axonal neurites elaborated during serum deprivation. To examine whether or not tau played an essential role in axonal outgrowth, cells cultured in serum-free medium were treated at 24 h intervals with antisense- and sense-oriented cDNA oligonucleotides (25 or 36 mers that span or are upstream of tau initiation codon) and were simultaneously serum deprived. Oligonucleotide uptake was confirmed by determination of intracellular levels of radiolabeled oligonucleotides. Treatment for 48 h with tau antisense oligonucleotides reversibly inhibited the expression of tau and the number of neurite-bearing cells compared with treatment with sense oligonucleotides. By contrast, tubulin expression was not affected. When cells were treated with antisense oligonucleotide simultaneously with serum deprivation, the initial outgrowth of neurites was unaffected, but continued neurite elongation was prevented. By contrast, neurite outgrowth at 4 h was inhibited when cells were pretreated with tau antisense 24 h before serum deprivation. Furthermore, intracellular delivery of anti-tau antiserum prevented neurite outgrowth and, in cells that had previously been deprived of serum for 24 h, induced retraction of existing neurites. These findings indicate that both the initiation and the continued outgrowth of neurites are dependent on tau and that pre-existing cytoplasmic pools of tau can mediate initial neuritogenesis.