Craniofacial Defects in Embryos with Homozygous Deletion of Eftud2 in Their Neural Crest Cells Are Not Rescued by Trp53 Deletion

Embryos with homozygous mutation of Eftud2 in their neural crest cells (Eftud2(ncc-/-)) have brain and craniofacial malformations, hyperactivation of the P53-pathway and die before birth. Treatment of Eftud2(ncc-/-) embryos with pifithrin-alpha, a P53-inhibitor, partly improved brain and craniofacial development. To uncover if craniofacial malformations and death were indeed due to P53 hyperactivation we generated embryos with homozygous loss of function mutations in both Eftud2 and Trp53 in the neural crest cells. We evaluated the molecular mechanism underlying craniofacial development in pifithrin-alpha-treated embryos and in Eftud2; Trp53 double homozygous (Eftud2(ncc-/-); Trp53(ncc-/-)) mutant embryos. Eftud2(ncc-/-) embryos that were treated with pifithrin-alpha or homozygous mutant for Trp53 in their neural crest cells showed reduced apoptosis in their neural tube and reduced P53-target activity. Furthermore, although the number of SOX10 positive cranial neural crest cells was increased in embryonic day (E) 9.0 Eftud2(ncc-/-); Trp53(ncc-/-) embryos compared to Eftud2(ncc-/-) mutants, brain and craniofacial development, and survival were not improved in double mutant embryos. Furthermore, mis-splicing of both P53-regulated transcripts, Mdm2 and Foxm1, and a P53-independent transcript, Synj2bp, was increased in the head of Eftud2(ncc-/-); Trp53(ncc-/-) embryos. While levels of Zmat3, a P53- regulated splicing factor, was similar to those of wild-type. Altogether, our data indicate that both P53-regulated and P53-independent pathways contribute to craniofacial malformations and death of Eftud2(ncc-/-) embryos.

Automated summary

This study demonstrates that craniofacial malformations and death in Eftud2(ncc-/-) embryos are caused by hyperactivation of the P53 pathway. Treatment with pifithrin-alpha partially improves craniofacial development, while Trp53 homozygous mutation reduces neural tube apoptosis and P53-target activity. However, the brain and craniofacial development remain unaffected in embryos with both Eftud2 and Trp53 mutations.