Epiblast Lumenogenesis is not a mammalian-specific trait

Epiblast lumenogenesis, which leads to amniotic cavity formation, is a hallmark of mammalian embryogenesis, and required for crucial developmental processes such as anterior-posterior patterning and gastrulation. Based on avian model-organisms, the epiblast in reptiles is thought to form a monolayered flat disc that undergoes anterior-posterior patterning and gastrulation. Here, we report that the squamate, veiled chameleon ( Chameleo calyptratus ), exhibits epiblast lumenogenesis and amniogenesis prior to anterior-posterior patterning. Using SEM, immunofluorescence, and histology techniques, we demonstrate that chameleon epiblast lumenogenesis occurs via a purse-string-like mechanism involving the formation of supracellular actin cables in concentric rings around the epiblast followed by constriction that closes the epiblast lumen. Through expression analyses of Nodal1 , Nodal2 , Cerberus , Lefty , Brachyury, Wnt3A , and Bmp2 , and immunostaining for Brachyury, we uncovered a Wnt3a- and Brachyury-positive ring at the edge of the epiblast concomitant with lumenogenesis, and preceding anterior-posterior patterning and gastrulation. Furthermore, we report anterior-posterior patterning in veiled chameleons occurs independently of Cerberus and Lefty . These processes that mediate epiblast lumenogenesis in chameleons, result in a morphology remarkably similar to human embryos, despite 300 million years of evolutionary separation. Taken together, we show that pre-gastrulation epiblast lumenogenesis is not mammalian-specific, but also occurs in some non-avian reptiles.