Differential requirement of ZIC3 function in cardiac development and X-linked heterotaxy

Heterotaxy, contributing to ~5% of congenial heart defects (CHD), arises from abnormal left-right patterning. Mutations of ZIC3 gene (Zinc finger protein of cerebellum 3) are associated with human Xlinked heterotaxy. A mouse model with targeted disruption of Zic3 exhibited ~75% early lethality, and recapitulated the phenotype seen in human patients. However, it is not knownwhether ZIC3 is required in a single developmental field or whether it has pleiotropic roles in multiple developmental processes, and the detailed mechanism remains elusive. To address these questions, we generated a conditional allele of the Zic3 gene by flanking its 1st exon with loxP sites. Sox2-cre, Wnt1-cre and T-cre lines were used to delete Zic3 in epiblast, neural crest and mesoderm, respectively. Deletion of Zic3 in epiblast and mesoderm, but not in neural crest, led to ~50% early lethality. Examination of epiblast conditional embryos by microscopy revealed multiple CNS and neural tube defects similar to the null embryos. But these defects were not found in mesoderm or neural crest conditional embryos, suggesting that Zic3's function in CNS development likely remains intact in these mutants. MRI scanning of Zic3 epiblast and mesoderm conditional embryos also uncovered multiple heterotaxy related visceral abnormalities. Gene expression analysis by microarray in the hearts of embryos at 15.5 dpc revealed a similar expression pattern between Zic3 epiblast conditional and null males, which was significantly different from control males. Perturbed expression of several cardiac genes and direct targets of Zic3 suggested that Notch, BMP and TGF-β signaling might be affected, and requires further investigation.