CpG Binding Protein Is Crucial for Early Embryonic Development

Abstract

Epigenetic modification of DNA via CpG methylation is essential for the proper regulation of gene expression during embryonic development. Methylation of CpG motifs results in gene repression, while CpG island-containing genes are maintained in an unmethylated state and are transcriptionally active. The molecular mechanisms involved in maintaining the hypomethylation of CpG islands remain unclear. The transcriptional activator CpG binding protein (CGBP) exhibits a unique binding specificity for DNA elements that contain unmethylated CpG motifs, which makes it a potential candidate for the regulation of CpG island-containing genes. In order to assess the global function of this protein, mice lacking CGBP were generated via homologous recombination. No viable mutant mice were identified, indicating that CGBP is required for murine development. Mutant embryos were also absent between 6.5 and 12.5 days postcoitum (dpc). Approximately, one-fourth of all implantation sites at 6.5 dpc appeared empty with no intact embryos present. However, histological examination of 6.5-dpc implantation sites revealed the presence of embryo remnants, indicating that CGBP mutant embryos die very early in development. In vitro blastocyst outgrowth assays revealed that CGBP-null blastocysts are viable and capable of hatching and forming both an inner cell mass and a trophectoderm. Therefore, CGBP plays a crucial role in embryo viability and peri-implantation development.

ACKNOWLEDGMENTS

We acknowledge David Williams (Herman B Wells Center for Pediatric Research, Indiana University, Indianapolis, Ind.) and Celeste Simon (University of Pennsylvania, Philadelphia) for helpful discussions regarding construction of the targeting vector. We also thank Joseph Ruiz and John Critser (Herman B Wells Center for Pediatric Research) for their generous assistance in analysis of the mutant embryos and Gen-Sheng Feng (Burnham Insitute, La Jolla, Calif.) for providing the pPNT vector. We also acknowledge LeAnn Boldridge and the Center for Excellence in Molecular Hematology for histological sectioning; the Indiana University Cancer Center mouse core facility for ES cell transfections, blastocyst injections, and generation of chimeric animals; and Paula D. Ladd, Angela Nevins, and Mark Starr for their technical assistance.

This work was supported by the Riley Memorial Association, Public Health Service grant CA58947 from the National Cancer Institute (D.G.S), an NRSA from the NIH (D.L.C.), and an American Heart Association postdoctoral fellowship (D.L.C.).