Deep Proteome of the Developing Chick Midbrain

The epithelial-to-mesenchymal transition (EMT) and migration
of
cranial neural crest cells within the midbrain are critical processes
that permit proper craniofacial patterning in the early embryo. Disruptions
in these processes not only impair development but also lead to various
diseases, underscoring the need for their detailed understanding at
the molecular level. The chick embryo has served historically as an
excellent model for human embryonic development, including cranial
neural crest cell EMT and migration. While these developmental events
have been characterized transcriptionally, studies at the protein
level have not been undertaken to date. Here, we applied mass spectrometry
(MS)-based proteomics to establish a deep proteomics profile of the
chick midbrain region during early embryonic development. Our proteomics
method combines optimal lysis conditions, offline fractionation, separation
on a nanopatterned stationary phase (μPAC) using nanoflow liquid
chromatography, and detection using quadrupole–ion trap–Orbitrap
tribrid high-resolution tandem MS. Identification of >5900 proteins
and >450 phosphoproteins in this study marks the deepest coverage
of the chick midbrain proteome to date. These proteins have known
roles in pathways related to neural crest cell EMT and migration such
as signaling, proteolysis/extracellular matrix remodeling, and transcriptional
regulation. This study offers valuable insight into important developmental
processes occurring in the midbrain region and demonstrates the utility
of proteomics for characterization of tissue microenvironments during
chick embryogenesis.