The overall goal of this proposal is to investigate the role of Cdx proteins in patterning the nascent neural crest (NC) along the body axis. The NC is a multipotent cell population that migrates extensively and gives rise to a remarkable array of cell types. While substantial evidence suggests that cranial and trunk NC cells (NCCs) exhibit differences in cellular behaviors, differentiation, and the underlying transcriptional network, the mechanism by which these differences are established remains unaddressed. The strategy proposed to investigate this key developmental question will be to focus on the role of Cdx, a homeodomain transcription factor, in patterning the NC along the anteroposterior (AP) axis. Cdx function will be studied in zebrafish embryos, as these are ideal for visualizing NCC behaviors in real time and well suited for genomic approaches. Moreover, the availability of powerful transgenic tools and sophisticated genetic approaches make it possible to label and modify specific cell types and evaluate gene function. Published and preliminary results have led to my central hypothesis that zebrafish Cdx proteins repress cranial NC identity, and promote the establishment of the trunk NC developmental program. Cdx proteins play a conserved role in AP patterning and have been shown to promote spinal cord identity and repress the hindbrain developmental program in the zebrafish. Given the role of Cdx proteins in patterning the neurepithelium, I propose them as candidates for establishing the differences between cranial and trunk NCCs. To evaluate this hypothesis, in Aim 1 I will determine whether Cdx regulates axial-specific NC features. In particular, I will determine whether loss and gain of Cdx function impacts axial-specific cellular behaviors and differentiation potential using Single Plane Illumination Microscopy and related approaches.
In Aim 2 I will fully characterize details of Cdx expression during NC development using fluorescent reporters, and utilize a novel approach to profile the genomic loci bound by Cdx4 in the NCC lineage. From this data, I expect to identify targets of direct regulation by Cdx4 and determine how Cdx establishes the trunk NC gene regulatory network. This project is innovative in using novel transgenic approaches, cutting-edge microscopy, and newly developed methods for profiling transcription factor-DNA interactions in order to investigate the role of Cdx in establishing trunk NCC identity, behaviors, and the underlying transcriptional network. The proposed research is significant as it addresses a long-standing and fundamental question in the field. Importantly, this project will provide an ideal training opportunity to support my long-term professional goal of becoming an independent academic investigator focused on investigating patterning and cell-fate regulation during development and developing innovative approaches and technologies that will enable scientific exploration.! !
A better understanding of neural crest cell development is critical for uncovering the mechanistic basis of related diseases, which account for some of the most common birth defects in live births and several highly metastatic cancers. A fundamental question in the field is how this remarkable multipotent cell population is patterned along the primary body axis, such that cells that arise in the head and the trunk exhibit marked differences in cellular behaviors, differentiation potential, and the underlying transcriptional network. In this proposal, I will evaluate the hypothesis that Cdx proteins, which have been previously shown to pattern the central nervous system along the body axis, regionalize the neural crest by promoting trunk neural crest identity and repressing the cranial neural crest developmental program. !
