Dissecting the Molecular Role of TBX1 in the Context of Human Pharyngeal Endoderm Development

PROJECT SUMMARY 22q11.2 Deletion Syndrome (22q11.2DS), the most common of the microdeletion syndromes, is caused by hemizygous loss of 0.7-3 Mb of DNA on chromosome 22 and results in a constellation of clinical phenotypes. The core phenotype originates from disrupted development of the pharyngeal apparatus. Particularly affected are the second heart field-dependent heart structures, great vessels, parathyroids, thymus, and lower craniofacial and face muscles. Although approximately 50 genes may be deleted, it is the haploinsufficiency of the transcription factor TBX1 that recapitulates most of the critical phenotype associated with 22q11.2DS. Genetic and developmental mouse studies have established that TBX1 is critical for typical development of the pharyngeal endoderm, a transient anatomical structure necessary for development of the thymus, parathyroids, and 4th pharyngeal arch arteries. Despite this central role, very little is known regarding the molecular mechanisms by which TBX1 functions in the pharyngeal endoderm. While a handful of studies have attempted to study the role of TBX1 in human cells, the cell types they have been conducted in are not representative of the appropriate developmental stage where and when TBX1 plays its critical role. To date, an effort to integrate all the critical genes into a pharyngeal endoderm or 4th pharyngeal arch arteries network has not been attempted, particularly in human cells. This R01 leverages recent development of an in vitro model which faithfully mimics the formation and progression of human pharyngeal endoderm, thereby providing an unprecedented opportunity to tease out the functions of TBX1 in its physiological context. Specifically, this model will be used to identify the transcriptional targets and partners of TBX1 (Aim1), investigate the role of TBX1 as epigenetic regulator of the human pharyngeal endoderm (Aim2), and mechanistically investigate newly discovered putative regulatory regions of the TBX1 locus (Aim3). The overarching hypothesis is that TBX1 is at the center of a Gene Regulatory Network critical for both the formation and maturation of the pharyngeal endoderm and the morpho-patterning of the surrounding mesoderm and neural crest cells. The proposed work is expected to identify the molecular mechanism at the basis of TBX1 haploinsufficiency and identify pathways which could be rescued through pharmacological intervention. Dissection of the epigenetic and molecular machinery responsible for pharyngeal endoderm formation will be instrumental in informing the generation of cell therapies for 22q11.2DS.