Abstract
Purpose: :
We aimed to generate a mouse model of human autosomal dominant optic atrophy which is caused by heterozygous mutation in the OPA1 gene (human 3q28/ mouse chromosome 16), and leads to blindness through the loss of retinal ganglion cells (RGCs).
Methods: :
An ENU mutagenised DNA archive from C3H mice was screened using heteroduplex temperature gradient capillary electrophoresis for mutations in mouse opa1. A heterozygous missense mutation in exon 8 coding for a C to T transition at 1051bp was detected. This mutation is predicted to result in protein truncation (Gln 285 to Stop: Q285X), resembling human disease causing mutations at aa 290: R290W and R290Q (c.868C>T and c.869G>T). Sperm were used (IVF with C57Bl/6) to generate heterozygous F1 hybrid founders. PCR genotyping, excluding rd1 (pdeB), and opa1 allele–specific genotyping were used to direct breeding with WT C57Bl/6.
Results: :
At F3 no opa1–/– homozygous animals have been detected, and together with smaller than expected litter size this is evidence for opa–/– being embryonic lethal. Examination of embryos shows early embryonic lethality. No gross systemic or neurological anomalies were identified when F2 opa1+/– and WT litter mates were screened using the SHIRPA test or on histology. No significant retinal pathology was identified by dilated fundal examination or on H& E retinal RGC counts. However, anti–opa1 antibody staining of retina revealed a reduction in opa1 expression. Functional visual phenotyping, using the optokinetic response, failed to highlight a significant difference between 6 month old opa1+/– and WT littermates, but large differences within the opa1+/– visual acuity scores suggest variable expression.
Conclusions: :
Our finding of opa1–/– embryonic lethality suggests a vital early developmental role for opa1 and suggests that the GTPase and dynamin–central regions are of key importance to the protein function. The absence of a gross retinal phenotype at 6 months of age, highlights the need to age this model for a retinal phenotype to manifest, mirroring the heterogeneous nature of this human condition.
Keywords: transgenics/knock-outs • ganglion cells • retinal degenerations: hereditary