Figure 1. Cep57 controls Fgf2-mediated bone development.

(A) Gene targeting approach used to mimic the CEP57^T patient mutation. Shown are the relevant portion of the murine Cep57 locus (top), the targeting vector (the 11-bp duplication is highlighted in red) with the recombined hypomorphic allele (middle), and the final Cep57^T allele following Cre-mediated excision of the neomycin (NEO) selection cassette. (B) Images of pups several hours after birth (arrowhead marks the short curly tail). Scale bars: 5 mm. (C) H&E-stained sagittal sections of 1-day-old pups. Arrowheads indicate the degree of spinal cord curvature. Scale bars: 1 mm. High magnification of the areas marked by red and green boxes are shown to the right. Scale bars: 100 μm. (D) Images of alizarin red– and Alcian blue–stained bone (red) and cartilage (blue) tissue of 1-day-old pups. Indicated are lumbar vertebra 4 (L4), sacral vertebra 1 (S1), and caudal vertebra 1 and 8 (Ct1 and Ct8) landmarks. Inset (yellow box) shows defective (bifid) vertebral body ossification. Scale bars: 1 mm. (E) Western blot analysis of brain and lung lysates of 1-day-old Cep57^+/+, Cep57^+/T, and Cep57^T/T mice. Ponceau (PonS) served as loading control. (F) Representative images of sagittal sections from thoracic and lumbosacral vertebral regions of 1-day-old pups immunostained for Fgf2. Arrowheads indicate regions with Fgf2 expression. Scale bars: 100 μm. (G) Analysis of Fgf2 subcellular localization using immunofluorescence in paraffin sections of the lumbosacral region of 1-day-old pups labeled for Fgf2. Nuclei were visualized with Hoechst. Scale bar: 5 μm. (H) Western blots of tissue lysates probed for Fgf2. Shown are the 35-kDa (full-length) and 18-kDa isoforms of Fgf2. PonS served as the loading control. Western blots are representative of 3 independent experiments.