The single neurofilament subunit of the lamprey forms filaments and regulates axonal caliber and neuronal size in vivo

Neurofilaments (NFs) are composed of a heteropolymer of three related subun its in mammalian neurons, where they are a major component of the cytoskele ton in large neurons and are thought to regulate axonal diameter. NFs in th e lamprey, while ultrastructurally and functionally indistinguishable from mammalian NFs, are polymers of a single subunit protein, NF180. In this stu dy, we use the simplicity of lamprey NFs and the accessibility of the lampr ey central nervous system (CNS) to examine the effects of overproducing NFs in an identified giant neuron in vivo, and thus to elucidate the role of N Fs in regulating neuronal size and axonal caliber in the vertebrate CNS. We show that overexpression of NF180 tagged with a variant of Green Fluoresce nt Protein (EYFP) in identified lamprey neurons (ABCs) and in human neurobl astoma (NB2a) cells results in the assembly of exogenous NF180 into ultrast ructurally normal NFs that are tightly packed and unphosphorylated. These a ccumulate in the somata of NB2a cells and produce somatic swelling by 3 day s post-transfection. NF180 overexpression in lamprey ABCs in vivo causes ex ogenous NFs to accumulate in ABC axons, somata, and dendrites, and induces a significant increase in axonal diameter without increasing axonal NF pack ing density. Overexpression of EYFP alone has none of these effects. We con clude that NF180 normally plays a critical role in determining axonal calib er in ABCs and may influence neuronal size in situations where NFs accumula te in the soma, such as after axonal injury. Cell Motil. Cytoskeleton 46: 1 66-182, 2000. (C) 2000 Wiley-Liss, Inc.