Analysing the role of the anterior visceral endoderm in anterior-posterior axis establishment in the mouse embryo

Abstract

Anterior-posterior (A-P) axis specification requires signalling from a specialised extraembryonic tissue called the anterior visceral endoderm (AVE). AVE precursors are induced at the distal tip of the pre-gastrulating mammalian embryo and move to the prospective anterior. This movement is essential in correctly positioning the A-P axis. Firstly, the contribution of proliferation to AVE movements has been tested. Previous studies have suggested that differential proliferation within the visceral endoderm (VE) is required to determine its direction by passive displacement. High-resolution imaging and embryo culture experiments have been used to assess the distribution and requirements of proliferation during AVE movements. Differential proliferation was not observed along the A-P axis of the VE. However, proliferation within the epiblast was found to be required for correct AVE movements in a Nodal-dependant manner. Secondly, to address the function of the AVE in vivo, this tissue was genetically ablated. Diphtheria toxin A was knocked-in to the Hex locus (an early AVE marker) by homologous recombination in embryonic stem cells. Depending on the time of ablation the AVE or its precursors could be ablated as demonstrated by AVE marker analysis. Ablation of the AVE precursors prior to 5.5 dpc resulted in embryos that displayed greatly disturbed patterning and were unable to gastrulate. AVE ablation between 5.5-6.5 dpc resulted in delayed formation of the primitive streak, epithelial to mesenchymal transition defects and a mispatterning of the anterior primitive streak. A decrease in Nodal signalling was found to be a likely cause for the defects observed in these ablated embryos, indicating that AVE maintains Nodal signalling levels in the epiblast. Together these experiments find a role for epiblast proliferation in AVE migration and identify a novel role for the AVE in patterning the primitive streak. Therefore this investigation provides a critical insight into how the A-P axis is established in mammals.