Aey17, a new small–eye phenotype caused by a mutation in Fgf10

Abstract

Abstract: : Purpose: Aim of the study was the phenotypic and genetic characterization of a new mouse mutant, Aey17, which was picked up by its small eyes. Methods: Aey17 was established as a new mutant line in the mouse; a genome–wide linkage analysis was performed using microsatellite markers. Genomic DNA was isolated from embryonic tails (E14.5). RNA was isolated from embryonic head (E14.5), transcribed into cDNA and used as template for PCR according to standard procedures. Results:Among offspring of C3HeB/FeJ mice treated with ethylnitrosourea (ENU), a new small–eye phenotype was identified. Slit–lamp examination demonstrated variable additional ocular anomalies like cornea opacities, glittering spots at the cornea or polar lens opacities. Homozygous mutants are not viable; some offspring were born without arms and legs. Genome–wide linkage analysis (after outcross/backcross to C57BL/6J mice) mapped the mutation on the telomer of mouse chromosome 13 close to the marker D13Mit35. Therefore, Fgf10 was tested as a candidate gene for the mutation. Sequence analysis of the Fgf10 transcript revealed an insertion of 49 bp at the beginning of exon 2. Since the causative mutation is an A–>G substitution destroying the canonical AG/GT splice site at the end of intron 1/beginning of exon 2, a similar sequence (AG/CT) 49 bp upstream is used in the mutants as new splice site leading to the observed insertion in the mature mRNA. On the translational level, it leads to a shift in the open reading frame, and the protein will be truncated after 54 new amino acids. In the mutant Fgf10 protein, the domain characterizing the entire family of heparin–binding growth factors (HBGF domain) is missing together with the majority of the phosphorylation sites. Conclusions:A new mouse mutant (Aey17) was characterized by a small–eye phenotype. Linkage and sequence analysis demonstrated a causative mutation in the Fgf10 gene. The ocular phenotype of the microphthalmic Aey17 mutants deviates from the phenotype reported in the Fgf10 knockout mutants described previously without effect on the eye size.

Keywords: gene mapping • genetics • growth factors/growth factor receptors