Disruption of type IV intermediate filament network in mice lacking the neurofilament medium and heavy subunits

To clarify the role of the neurofilament (NF) medium (NF-M) and heavy (NF-H ) subunits, we generated mice with targeted disruption of both NF-M and NF- H genes. The absence of the NF-M subunit resulted in a two- to threefold re duction in the caliber of large myelinated axons, whereas the lack of NF-H subunits had little effect on the radial growth of motor axons. In NF-M-/- mice, the velocity of axonal transport of NF light (NF-L) and NF-H proteins was increased by about twofold, whereas the steady-stale levels of assembl ed NF-L were reduced. Although the NF-M or NF-H subunits are each dispensab le for the formation of intermediate filaments, the absence of both subunit s in double NF-M; NF-H knockout mice led to a scarcity of intermediate fila ment structures in axons and to a marked approximately twofold increase in the number of microtubules. Protein analysis indicated that the levels of N F-L and alpha-internexin proteins were reduced dramatically throughout the nervous system. Immunohistochemistry of spinal cord from the NF-M-/-;NF-H-/ - mice revealed enhanced NF-L staining in the perikaryon of motor neurons b ut a weak NF-L staining in axons. In addition, axonal transport studies car ried out by the injection of [S-35]methionine into spinal cord revealed aft er 30 days very low levels of newly synthesized NF-L proteins in the sciati c nerve of NF-M-/-;NF-H-/- mice. The combined results demonstrate a require ment of the high-molecular-weight subunits for the assembly of type IV inte rmediate filament proteins and for the efficient translocation of NF-L prot eins into the axonal compartment.