Tuesday, May 14, 2024
5/14/2024
PROJECT SUMMARY: Hemizygous microdeletions encompassing TBX1 cause 22q11.2 deletion syndrome (22q11.2DS), the most common deletion syndrome in humans. Among a broad spectrum of clinical features that often include craniofacial dysmorphia, congenital heart defects (CHDs) of the outflow tract (OFT), and aortic arch, which is derived from embryonic vessels termed pharyngeal arch arteries (PAAs), are the main causes of mortality during childhood. Importantly, specific craniofacial muscles, the OFT, and PAAs all derive from the pharyngeal apparatus, a transient series of arches and pouches that form by segmentation on the lateral surface of the head. Because segmentation is severely compromised in Tbx1-deficient animals, their craniofacial and cardiovascular defects were considered secondary. Using zebrafish as a model, my laboratory reported that tbx1 mutant embryos recapitulate several aspects of 22q11.2DS, including craniofacial muscle, OFT, and PAA deficiencies. We also demonstrated that these end structures descend from nkx2.5+ progenitors initially specified in the anterior lateral plate mesoderm (ALPM) before becoming sequestered in the cores of the pharyngeal arches. Unexpectedly, we discovered that tbx1 mutants fail to specify the nkx2.5+ pharyngeal lineage in the ALPM that gives rise to the missing derivatives. Because this phenotype precedes pharyngeal segmentation, defects in progenitor specification are likely causal for the deficiencies observed in tbx1 mutants. The aberrant fates of these progenitors, which maintain hand2 and gata5 expression in the ALPM, are unknown. Moreover, the molecular mechanisms by which Tbx1 directs specification of the pharyngeal lineage in the ALPM have not been elucidated in any model system. Here, we have the unique opportunity to uncover the molecular mechanisms by which Tbx1 directs specification of the pharyngeal progenitor cell lineage because we have defined the relevant developmental time window for analysis, identified markers of the preserved (gata5+) and missing (nkx2.5+) progenitor populations, implemented single-cell RNA and ATAC sequencing that allows for unprecedented resolution of changes in cellular populations and chromatin accessibility, and generated state-of-the-art genetic tools for tracking the aberrant fates of tbx1-expressing cells in tbx1 mutants and for identifying Tbx1 target loci that confer the pharyngeal progenitor cell fate. Overall, we propose to use the attributes of the zebrafish system, including its genetic tractability, unmatched embryonic accessibility, and imaging capabilities, to reveal new mechanistic insights that will provide the most comprehensive view of how Tbx1 is necessary and potentially sufficient for instructing the pharyngeal progenitor cell lineage during vertebrate development.
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