Abstract
Purpose:
Meckel syndrome is a lethal form of syndromic ciliopathy with pleiotropic clinical features like retinal dystrophy, mental retardation, polydactyly. The patients with mutations in CC2D2A (Coiled-Coil & C2 Domain containing 2A) displayed Meckel syndrome. CC2D2A protein is localized to the cilia basal body. The patient derived fibroblasts have basal body, however, they failed to develop cilia indicating a mechanistic disconnect in the basal body due to CC2D2A mutation. To examine this disconnect we have generated Cc2d2a loss of function diseases models in the mouse.
Methods:
The Cc2d2a-knockout (Cc2d2a-/-) mouse was generated by targeted deletion of exons 6 to 8 relying on homologous recombination. Immunochemistry and electron microscopy were employed to examine the basal body from mice fibroblasts.
Results:
The Cc2d2a-/- mouse is embryonic lethal. It displays developmental defects like situs inversus, heterotaxy, polydactyly, anophthalmia, hydrocephalus and liver fibrosis. Occasionally, the Cc2d2a-/- mouse survived for a month but was with severe hydrocephalus and retinal dystrophy. Analysis of the embryonic node, embryonic fibroblasts, and kidney tubules showed that cilia biogenesis was disrupted in the Cc2d2a-/- mouse. Embryonic fibroblasts (MEFs) isolated from Cc2d2a-/- mice lack cilia even though contain mother centriole (MC or basal body) and pericentriolar proteins. In addition, MEFs have reduced levels of Odf2 (associated with subdistal appendages (SDA)) and ninein. Transmission electron microscopy revealed lack of SDAs in Cc2d2a-/- MEFs and consistent with this observation immuno-EM showed Cc2d2a localization onto SDA.
Conclusions:
Our studies show that Cc2d2a function is critical for embryonic development. We conclude that Cc2d2a is essential for the assembly of SDA, which is anchoring cytoplasmic microtubules and prime MC for axoneme biogenesis.
Keywords: 740 transgenics/knock-outs •
698 retinal development •
597 microscopy: electron microscopy