Figure 6.

Effects of combined deletion of NF-H and NF-M on α-internexin and NF-L transport and levels in optic axons. hm-dko mice were generated by crossbreeding and screening. A–D, Mouse genomic DNA was screened for targeted disruption of both NF-H (A, B) and NF-M (C, D) homozygous mice through genotyping the NF-H and NF-M loci by hybridizing with a sequence from NF-H or NF-M. NF-H, NF-M, and α-internexin (α-int) were identified by immunoblots of total optic nerve protein (10–20 μg) from 3- to 4-month-old mice using mAbs to NF-H (N52), NF-M (NN18), and α-internexin (MAB5224), respectively. E–G, In mice lacking NF-H and NF-M (E, F), α-internexin levels are <10% of wild-type (wt) levels detected by immunoblotting (G), and NF-L proteins are barely detectable as shown previously (Yuan et al., 2003). H, I, Slow axonal transport analyses (7 d after injection) in wild-type control (H) and hm-dko mice (I), as in Figure 5, reveal barely detectable transport of α-internexin in hm-dko axons compared with wild-type mice. The positions of cytoskeletal proteins are indicated, and the nerve segments are numbered below the gels consecutively from the level of the eye. J, K, 2D transport analyses (7 d after injection) confirm dramatically reduced transport of α-internexin into hm-dko axons. The asterisks in J and K indicate Hsc70.