Neuroplastin gene schematic and alleles.

(A) Schematic of the wild type Neuroplastin gene locus consisting of eight exons that encode: three extracellular Ig domains; a transmembrane region (TM); and, a short intracellular region. Alternative splicing involving the inclusion, or skipping, of exon 2 leads to the production of transcripts encoding the larger Np65, or smaller Np55, protein isoform, respectively. Previously reported mutant alleles audio-1 [13] and pitch [12] are point mutations present in exons 3 and 6, respectively, affecting both isoforms. (B) the Nptn^{tm1a(EUCOMM)Hmgu} (Nptn^tm1a) is a knockout first, conditional-ready allele, that was used to generate both the Nptn^{tm1b(EUCOMM)Hmgu} (Nptn^tm1b) knockout allele lacking exons 5 and 6, and the Nptn^{tm1c(EUCOMM)Hmgu} (Nptn^fl) floxed allele. Subsequent exposure of the Nptn^fl allele to cre recombinase excises exons 5 and 6, resulting in a null allele (Nptn^tm1d) in cells expressing cre. (C) An isoform-specific Np65 knockout mutant (Nptn^Δexon2) was generated using CRISPR/Cas9-mediated deletion of 941 nucleotides encompassing exon 2. Mice homozygous for the Nptn^Δexon2 are unable to express the larger Np65 isoform, while expression of Np55 remains.