The role of the transcriptional repressor ZEB2 in human neural crest cell formation and craniofacial pathology

Project Summary
The objective of this proposal is to delineate the role of the transcriptional repressor ZEB2 during human neural
crest cell formation, craniofacial differentiation, and Mowat-Wilson Syndrome. The neural crest is a multipotent
embryonic cell population unique to vertebrates which migrates extensively and differentiates into a variety of
derivatives, including most of the craniofacial bone, cartilage, and dental organ tissues. Numerous human
pathologies are associated with abnormal neural crest cell development, including the recently delineated
neurocristopathy Mowat-Wilson Syndrome (MWS). MWS is a rare syndrome caused by de novo heterozygous
mutations in ZEB2 leading to a distinct facial phenotype, dental abnormalities, intellectual disability, and other
anomalies including Hirschprung disease which are indicative of neural crest cell defects at multiple axial levels.
However, significant genotypic and phenotypic variability has been observed among MWS patients, and the
underlying mechanisms that contribute to this disease are not well understood. Furthermore, while ZEB2 has
been implicated as a major developmental regulator of the nervous system as well as in neural crest cell
epithelial-to-mesenchymal transition and migration, recent evidence in our human neural crest cell model
indicates a crucial role for ZEB2 in the early formation of the neural plate border and neural crest. ZEB2 is known
to negatively regulate TGFb superfamily signaling, whose modulation is essential for proper neural crest
formation and differentiation. Therefore, it is hypothesized that ZEB2 functions reiteratively to establish the
repressive epigenetic landscape throughout neural crest cell formation and craniofacial development by
modulating proper TGFb superfamily signaling, and that the failure of this regulation underscores the MWS
pathology. To test this hypothesis, three specific aims are proposed. Aim 1 will examine the modular functionality
of ZEB2 in regulating the proper levels of TGFb signaling throughout wild-type human cranial neural crest cell
formation. In particular, the cis-regulatory interactions between ZEB2 and TGFb receptor activated Smad
proteins and the functional requirement of co-repressors leading to changes in chromatin accessibility will be
delineated. In Aim 2, the MWS pathology will be addressed through the establishment of induced pluripotent
stem cells from MWS patients of diverse genotypes. Using these tools, the precise transcriptomic and epigenetic
misregulation during MWS-based neural crest cell formation will be interrogated, providing much needed
molecular insight into this understudied pathology. In Aim 3 of this proposal, the role of ZEB2 in wild-type and
MWS human tooth formation will be interrogated through the establishment of a human neural crest cell-based
tooth organoid model. These results will reveal for the first time the molecular role of this transcriptional repressor
throughout multiple stages of human neural crest cell formation, differentiation, and pathology.

Grant Number: 5R00DE029878-05
NIH Institute/Center: NIH
Principal Investigator: Rebekah Charney