The triple C-type isoform knockout (TCKO) allele was generated by deleting the three variable exons (Figures 1A–B and S1A), which specifically removes the C-type genes without affecting the splicing of the remaining 19 A-type and B-type Pcdhg variable exons (see below). Pcdhgtcko/tcko mutants are born alive at the normal Mendelian ratio, but invariably die during the first day after birth. The mutant mice are readily distinguishable from wild type and heterozygous littermates by a characteristic hunched posture and limb tremors, as well as by severely compromised voluntary movements and reflexes (Figure 1C and Movie S1). Remarkably, these phenotypes are identical to those described for the Pcdhg full cluster deletion mice (Figure 1D and Movie S1), in which all Pcdhg isoforms are abolished (Wang et al., 2002b). In addition to the common phenotypes described above, we found that both lines of mutants also exhibit intense muscle stiffness (Figures 1F–F”) and umbilical hernia (Figures 1G–G”). Interestingly, these phenotypes closely resemble those of mutant mice deficient in VGAT (Wojcik et al., 2006), GAD67 (Asada et al., 1997), and Gephyrin (Feng et al., 1998), which are essential components for GABA and glycine production and transmission.
