Cellular analysis of cleavage-stage chick embryos reveals hidden conservation in vertebrate early development

Abstract

Birds and mammals, phylogenetically close amniotes with similar post-gastrula development, exhibit little conservation in their post-fertilization cleavage patterns. Data from the mouse suggest that cellular morphogenesis and molecular signaling at the cleavage stage play important roles in lineage specification at later (blastula and gastrula) stages. Very little is known, however, about cleavage-stage chick embryos, owing to their poor accessibility. This period of chick development takes place before egg-laying and encompasses several fundamental processes of avian embryology, including zygotic gene activation (ZGA) and blastoderm cell-layer increase. We have carried out morphological and cellular analyses of cleavage-stage chick embryos covering the first half of pre-ovipositional development, from Eyal-Giladi and Kochav stage (EGK-) I to EGK-V. Scanning electron microscopy revealed remarkable subcellular details of blastomere cellularization and subgerminal cavity formation. Phosphorylated RNA polymerase II immunostaining showed that ZGA in the chick starts at early EGK-III during the 7th to 8th nuclear division cycle, comparable with the time reported for other yolk-rich vertebrates (e.g. zebrafish and Xenopus). The increase in the number of cell layers after EGK-III is not a direct consequence of oriented cell division. Finally, we present evidence that, as in the zebrafish embryo, a yolk syncytial layer is formed in the avian embryo after EGK-V. Our data suggest that several fundamental features of cleavage-stage development in birds resemble those in yolk-rich anamniote species, revealing conservation in vertebrate early development. Whether this conservation lends morphogenetic support to the anamniote-to-amniote transition in evolution or reflects developmental plasticity in convergent evolution awaits further investigation.

DAZLExpression Explains Origin and Central Formation of Primordial Germ Cells in Chickens

“…Because the unubiquitous GFP expression could be driven by the CMV promoter, we also investigated p-pol II expression during the intrauterine stages and obtained similar results as reported in our previous study [52]. These data suggest that ZGA in chicken first occurs in the central cells between EGK stages II and III, and then expands toward the periphery; it does not occur simultaneously in all cells.…”

Vertebrate Embryonic Cleavage Pattern Determination

“…Another notable feature of these embryos is that the point at which the first two cleavage furrows meet is not centered at the middle of the embryo (Sheng 2014; Nagai et al 2015). There is a known correlation between asymmetric inheritance of maternally deposited factors and establishment of PGCs, but whether this is associated with off-center early cleavages is unclear.…”

“…As cleavage progresses, the embryo will reach 5–6 layers of cells in thickness before thinning again during and after gastrulation (Sheng 2014). A study found that approximately 75 % of surface cells (i.e., those in the uppermost layer) divided in a direction parallel to the blastoderm plane, yielding two daughters in the same layer, while deeper cells show more variation, with approximately 56 % dividing in a direction 30–90° from that of the blastoderm plane (Nagai et al 2015; see below). These data suggest that cleavage orientation only partly explains increasing layer number, but other mechanisms, such as rearrangement of already cellularized blastomeres, may also be involved.…”

The avian-specific small heat shock protein HSP25 is a constitutive protector against environmental stresses during blastoderm dormancy

“…Additionally, developmentally constitutive HSP30 homologues in Xenopus were first detected in early and mid-tailbud embryos, after gastrulation, but not soon after ZGA, which in Xenopus occurs at the 128- to 256-cell stage7. It was recently discovered that ZGA starts between EGK stages II and III in chicken3233. HSP25 and HSP30CL transcripts were also not induced immediately after ZGA, at EGK III, but earlier than the homologues in Xenopus .…”