Recent insights into the morphological diversity in the amniote primary and secondary palates.

The assembly of the upper jaw is a pivotal moment in the embryonic development of amniotes. The upper jaw forms from the fusion of the maxillary, medial nasal, and lateral nasal prominences, resulting in an intact upper lip/beak and nasal cavities; together called the primary palate. This process of fusion requires a balance of proper facial prominence shape and positioning to avoid craniofacial clefting, whilst still accommodating the vast phenotypic diversity of adult amniotes. As such, variation in craniofacial ontogeny is not tolerated beyond certain bounds. For clarity, we discuss primary palatogenesis of amniotes into in two categories, according to whether the nasal and oral cavities remain connected throughout ontogeny or not. The transient separation of these cavities occurs in mammals and crocodilians, while remaining connected in birds, turtles and squamates. In the latter group, the craniofacial prominences fuse around a persistent choanal groove that connects the nasal and oral cavities. Subsequently, all lineages except for turtles, develop a secondary palate that ultimately completely or partially separates oral and nasal cavities. Here, we review the shared, early developmental events and highlight the points at which development diverges in both primary and secondary palate formation.

Diversity in primary palate ontogeny of amniotes revealed with 3D imaging.

The amniote primary palate encompasses the upper lip and the nasal cavities. During embryonic development, the primary palate forms from the fusion of the maxillary, medial nasal and lateral nasal prominences. In mammals, as the primary palate fuses, the nasal and oral cavities become completely separated. Subsequently, the tissue demarcating the future internal nares (choanae) thins and becomes the bucconasal membrane, which eventually ruptures and allows for the essential connection of the oral and nasal cavities to form. In reptiles (including birds), the other major amniote group, primary palate ontogeny is poorly studied with respect to prominence fusion, especially the formation of a bucconasal membrane. Using 3D optical projection tomography, we found that the prominences that initiate primary palate formation are similar between mammals and crocodilians but distinct from turtles and lizards, which are in turn similar to each other. Chickens are distinct from all non-avian lineages and instead resemble human embryos in this aspect. The majority of reptiles maintain a communication between the oral and nasal cavities via the choanae during primary palate formation. However, crocodiles appear to have a transient separation between the oral and nasal cavities. Furthermore, the three lizard species examined here, exhibit temporary closure of their external nares via fusion of the lateral nasal prominences with the frontonasal mass, subsequently reopening them just before hatching. The mechanism of the persistent choanal opening was examined in chicken embryos. The mesenchyme posterior/dorsal to the choana had a significant decline in proliferation index, whereas the mesenchyme of the facial processes remained high. This differential proliferation allows the choana to form a channel between the oral and nasal cavities as the facial prominences grow and fuse around it. Our data show that primary palate ontogeny has been modified extensively to support the array of morphological diversity that has evolved among amniotes.

Divergent palate morphology in turtles and birds correlates with differences in proliferation and BMP2 expression during embryonic development.

During embryonic development, amniotes typically form outgrowths from the medial sides of the maxillary prominences called palatal shelves or palatine processes. In mammals the shelves fuse in the midline and form a bony hard palate that completely separates the nasal and oral cavities. In birds and lizards, palatine processes develop but remain unfused, leaving a natural cleft. Adult turtles do not possess palatine processes and unlike other amniotes, the internal nares open into the oral cavity. Here we investigate craniofacial ontogeny in the turtle, Emydura subglobosa to determine whether vestigial palatine processes develop and subsequently regress, or whether development fails entirely. We found that the primary palate in turtles develops similarly to other amniotes, but secondary palate ontogeny diverges. Using histology, cellular dynamics and in situ hybridization we found no evidence of palatine process development at any time during ontogeny of the face in the turtle. Furthermore, detailed comparisons with chicken embryos (the model organism most closely related to turtles from a molecular phylogeny perspective), we identified differences in proliferation and gene expression patterns that correlate with the differences in palate morphology. We propose that, in turtles, palatine process outgrowth is never initiated due to a lack of mesenchymal bone morphogenetic protein 2 (BMP2) expression in the maxillary mesenchyme, which in turn fails to induce the relatively higher cellular proliferation required for medial tissue outgrowth. It is likely that these differences between turtles and birds arose after the divergence of the lineage leading to modern turtles.