Lamprey neurofilaments combine in one subunit the features of each mammalian NF triplet protein but are highly phosphorylated only in large axons.
Compared with heteropolymeric assemblies of neurofilament (NF) triplet proteins in mammalian NFs, lamprey (Petromyzon marinus) NFs are homopolymers of 180 kDa subunits (NF180). We describe unique features of lamprey NF180 that distinguish it as a prototype of vertebrate NF subunits. These features may underlie key functions subserved by the earliest vertebrate NFs. Lamprey NF180 displays properties common to all intermediate filament (IF) proteins, but it also exhibits features that distinguish the mammalian triplet of NF subunits from all other IF proteins. For example, digestion of lamprey NF180 with chymotrypsin produces an insoluble 40 kDa core unit and releases a soluble fragment intermediate in size (140 kDa) to the carboxy-terminal (sidearm) extensions of the 2 high-molecular-weight (Mr) mammalian NF subunits. The core unit contains epitopes similar to those in the core of each mammalian NF triplet protein, while the soluble fragment contains other determinants similar to those in the sidearms of the 2 high-Mr mammalian NF polypeptides. Like these polypeptides, the immunological properties of some NF180 peripheral determinants were strongly affected by their phosphorylation state. Indeed, NF180 shares immunological similarities with the multiphosphorylation repeat domains in the high-Mr mammalian NF subunits. Further similarities with mammalian NF proteins include the preferential expression of poorly phosphorylated NF180 isoforms and of phosphate-dependent NF180 epitopes in axons of all sizes, and the restriction of nonphosphorylated NF180 isoforms to neuronal perikarya. In marked contrast to mammals, however, the most heavily phosphorylated isoforms of NF180 were expressed exclusively in large-diameter axons. We conclude that the single subunit forming lamprey NFs exhibits the essential features of mammalian NFs, i.e., a filament-forming core and a carboxy-terminal extension with a multiphosphorylation site. Further, the sharp restriction of heavily phosphorylated NF180 to large axons suggests that multiphosphorylation domains were acquired during evolution to permit larger axon diameters and faster conduction velocities.