Figure 5. Schwann cell neurofascin (NFasc) expression is abolished in the TDP-43 conditional knockout (cKO).
(A) The schema of mouse Nfasc locus showing the region from exon 13 to exon 33. The coding strand of Nfasc is located on the reverse strand of chromosome 1, as defined in the GRCm38/mm10 mouse genome assembly, and flipped here for reader convenience. Alternative exons specific to NF155 (exons 23 and 24) or NF186 (exons 28–30) are shown in blue while other alternative exons (exons 18 and 27) are shown in gray. NF140 contains neither exons 23 and 24 nor exons 28–30. The open rectangle denotes the untranslated region, and the solid ones the open reading frame. (B) Western blotting of P28 sciatic nerve homogenates probed for NFasc, contactin-associated protein (Caspr), and actin. (C) Immunostaining of P3 sciatic nerves for NFasc (green) and ankyrinG (AnkG) (magenta) showing pairs of approaching heminodes (nodes flanked by myelin sheaths only on one side). The paranodes are indicated by arrowheads. Scale bar, 5 μm. (D) RT-qPCR analysis of P28–P29 sciatic nerves shows the transcript levels relative to those of the wild-type (WT) after normalized to the internal control Polr2a. Exon 3 of Tardbp is the floxed exon, and the primer pair for Tardbp ex3 detects TDP-43 mRNA transcribed from the unrecombined allele. The primer pair for Tardbp exons 4–5 detects the nonfloxed region to confirm mRNA degradation after Cre recombination. Pan-Nfasc is detected by primers spanning exons 12–13, NF155 by primers spanning exons 22–23, NF140+ex27 by primers annealing to the junctions of exons 22/25 and exons 27/31, and NF140-ex27 by primers annealing to the junctions of exons 22/25 and exons 26/31. Bars represent mean ± SEM. n = 3 mice per genotype. *p<0.05; **p<0.01; ***p<0.001; ns, p≥0.05; one-sample unpaired two-tailed t-tests (WT vs. conditional heterozygote [cHet] and WT vs. cKO). cHet and cKO by Dhh-Cre (B–D).
Figure 5—figure supplement 1. Localization of neurofascin (NFasc) in the Schmidt–Lanterman incisure is not affected in the contactin-associated protein knockout (Caspr KO).
