Unexpected CEP290 mRNA splicing in a humanized knock-in mouse model for Leber congenital amaurosis.
Leber congenital amaurosis (LCA) is the most severe form of retinal dystrophy with an onset in the first year of life. The most frequent genetic cause of LCA, accounting for up to 15% of all LCA cases in Europe and North-America, is a mutation (c.2991+1655AG) in intron 26 of CEP290. This mutation generates a cryptic splice donor site resulting in the insertion of an aberrant exon (exon X) containing a premature stop codon to CEP290 mRNA. In order to study the pathophysiology of the intronic CEP290 mutation, we generated two humanized knock-in mouse models each carrying ~6.3 kb of the human CEP290 gene, either with or without the intronic mutation. Transcriptional characterization of these mouse models revealed an unexpected splice pattern of CEP290 mRNA, especially in the retina. In both models, a new cryptic exon (coined exon Y) was identified in ~5 to 12% of all Cep290 transcripts. This exon Y was expressed in all murine tissues analyzed but not detected in human retina or fibroblasts of LCA patients. In addition, exon x that is characteristic of LCA in humans, was expressed at only very low levels in the retina of the LCA mouse model. Western blot and immunohistochemical analyses did not reveal any differences between the two transgenic models and wild-type mice. Together, our results show clear differences in the recognition of splice sites between mice and humans, and emphasize that care is warranted when generating animal models for human genetic diseases caused by splice mutations.
Department of Human Genetics, Radboud University Medical Centre, Nijmegen, The Netherlands ; Institute for Genetic and Metabolic Disease, Radboud University Medical Centre, Nijmegen, The Netherlands.
Alejandro
A
Garanto
Sylvia E C
SE
van Beersum
Theo A
TA
Peters
Ronald
R
Roepman
Frans P M
FP
Cremers
Rob W J
RW
Collin
D000818:Animals; D000951:Antigens, Neoplasm / Q000235:genetics*; D018797:Cell Cycle Proteins; D002460:Cell Line; D003598:Cytoskeletal Proteins; D004195:Disease Models, Animal; D005091:Exons / Q000235:genetics; D005260:Female; D055879:Gene Knock-In Techniques; D006801:Humans; D057130:Leber Congenital Amaurosis / Q000235:genetics*; D051379:Mice; D009363:Neoplasm Proteins / Q000235:genetics*; D011247:Pregnancy; D012326:RNA Splicing; D012160:Retina / Q000378:metabolism; D013045:Species Specificity; D014158:Transcription, Genetic / Q000235:genetics
D000951:Antigens, Neoplasm; D018797:Cell Cycle Proteins; C493903:Cep290 protein, human; D003598:Cytoskeletal Proteins; D009363:Neoplasm Proteins