Project Summary/Abstract
The neural crest and ectodermal placodes, unique and defining features of the vertebrate embryo, arise from
the region of the ectoderm termed the neural plate border, which is juxtaposed between the neural plate and the
non-neural ectoderm of the gastrula. They constitute the origins of the craniofacial structures as well as the
cranial ganglia, in addition to other diverse cell and tissue types. Malformation of these cells or their derivatives
can have devastating effects on embryonic development, resulting in pathologies including craniofacial
syndromes, congenital heart defects, and cancers. Nonetheless, exceedingly little is known of the genetic and
epigenetic mechaniasms by which neural crest and ectodermal placode cells become specified. The goal of this
proposal is to investigate the epigenetic regulation of micro RNAs (miRNAs) during specification of neural plate
border cells in order to understand how this diverse pool of progenitors becomes differentially primed for the
acquisition of distinct fates. Importantly, we will address this question in the context of developmentally critical
vitamins, folate and Vitamin C, which are essential cofactors for DNA methylation and demethylation upstream
of miRNA expression. We will combine advanced sequencing technologies with classical techniques in
embryology to test the hypothesis that Vitamin C and folate are required for establishing proper DNA
methylation patterns on miRNA regulatory elements to prime neural plate border cells for fate
specification. Specifically, we will (1) perform transcriptome-wide profiling of miRNAs in progenitors of the
neural plate and neural plate border at different developmental timepoints to uncover spatiotemporal miRNA
signatures underlying differential specification, (2) perform scATAC-seq to identify differential chromatin
accessibilty (i.e. putative regulatory elements), and perform oxBS-seq to reveal differential mehtyation states of
these elements, and (3) determine the respective roles of the essential DNA (de)methylation cofactors folate and
Vitamin C in regulating methylation states on miRNA regulatory elements during fate specifcation. This proposal
seeks to provide the first transcriptome-wide analysis of miRNAs and the corresponding mRNA targets that they
regulate, and the first comprehensive assessment of chromatin landscape (i.e. chromatin accessibility revealed
by ATAC-seq and 5hmC/5mC revealed by oxBS-seq) in neural plate and neural plate border tissues. Moreover,
results from the proposed experiments will contribute fundamental knowledge of the complex mechanisms
regulating fate specification at the neural plate border. Our findings will also have important implications for the
understanding of birth defects arising from malformation of the neural tube, neural crest, and ectodermal
placodes. The proposed project will be carried out under the guidance of Dr. Marianne Bronner who is a leader
in the field of developmental biology and an expert in neural crest development. She is the director of Caltech’s
Beckman Institute where her lab has access to outstanding core facilities (e.g. advanced microscopy and
sequencing centers) that will be instrumental to the successful completion of the proposed study.