Tracheal Separation is Driven by NKX2-1-Mediated Repression of <i>Efnb2</i> and Regulation of Endodermal Cell Sorting

The mechanisms coupling fate specification of tissues to their physical separation remain largely unknown. mammalian trachea and esophagus differentiate from a single tube definitive endoderm, requiring the mutually repressive transcription factors SOX2 NKX2-1, but how dorsoventral site tissue is defined properly allocate tracheal esophageal cell types Here, we show that EPH/EPHRIN signaling gene Efnb2 regulates tracheoesophageal by controlling allocation tracheal-fated cells. Ventral loss NKX2-1 results in disruption an expansion expression independent SOX2. Through chromatin immunoprecipitation, find binds gene, indicating repression likely direct. Finally, inducing mosaicism for Nkx2-1 within embryonic foregut, generate ectopic NKX2-1/EPHRIN-B2 boundaries which organize separation. Together these data demonstrate connects with through regulation EPHRIN-B2 sorting.