Developmental abnormalities of glycosylphosphatidylinositol-anchor-deficient embryos revealed by Cre/IoxP system

One mode used to link membrane proteins to a cell membrane is by means of a special glycolipid anchor termed glycosylphosphatidylinositol (GPI). Pig-a , an X-linked gene, is involved in the first step of GPI-anchor biosynthesi s. Disruption of this gene causes cessation of GPI biosynthesis on the endo plasmic reticulum, thereby leading to the absence of GPI-anchored proteins on the cell surface. We have previously reported that mice with high chimer ism was never obtained from Pig-a disrupted ES cells, suggesting that GPI-a nchored protein(s) may have important roles for mouse development such that the absence of GPI-anchored proteins causes a lethal effect to mice. In th is study, this lethal effect: has been investigated by using a conditional approach to "knockout" the Pig-a gene. For this, mice harboring a Pig-a gen e flanked by two loxP sites (Pig-a(flox)) were mated with hCMV-Cre transgen ic mice, which express Cre recombinase before implantation. The allele disr uptions were identified by PCR analysis of embryo yolk sac DNA. Embryos har boring a complete disruption of Pig-a gene ceased to develop beyond the nin th day of gestation. Female embryos in which one Pig-a allele was disrupted by Cre such that only half of the cells in the embryo proper did not expre ss GPI-anchored proteins due to random X inactivation developed until 19 da ys post coitum (dpc), but showed abnormal phenotypes such as insufficient c losure of neural tube and cleft palate. These data further highlight the im portance of GPI-anchored proteins during mouse embryonic development.