Molecular Interactions During Neural Crest Formation

Yale University, New Haven CT

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Abstract

DESCRIPTION (provided by applicant): Neural crest stem cells are involved in human illnesses including tumors and craniofacial and heart malformations. They generate most of the cells of the peripheral nervous system, craniofacial bone and cartilage, cells of the outflow tract of the heart and melanocytes amongst other derivatives. Studying the basic developmental biology of the neural crest is a critical step to better understand, diagnose, and treat these human conditions. The long term objective of this proposal is to study the molecular interactions at play during neural crest cell formation at the earliest possible stages. Therefore it is imperative to identify the time, place and cells or tissues involved in their induction. It has been recently shown that a region of the epiblast is specified to form neural crest cells during gastrulation, these same region will express Pax7 soon after, and that Pax7 is required for neural crest formation. In light of these findings it is proposed to investigate when, where and how neural crest cells are induced and specified though a combination of classic embryology and molecular biology approaches. The specific aims are: 1) identify time, location and tissues involved in the induction and specification of the neural crest;2) characterize the participation of BMP, FGF and WNT molecules in neural crest formation;and 3) Characterize Pax7 molecular interactions during neural crest formation. These aims are guided by a set of testable hypotheses that analyze neural crest cell development at the blastula/gastrula stage, challenge the accepted cannon of their induction through neural and non-neural/mesodermal interactions, and analyze at the molecular level the participation of three signaling pathways and the mode of action of Pax7. In vivo and in vitro experiments will record the progression of neural crest developments with a battery of molecular markers. To this end, specification and induction maps will be obtained through collagen explants and grafts to naive embryonic environments from epiblast tissues to be tested (specific aim 1);distinct BMP, FGF and WNT signaling conditions (including combinatorial activation and inhibition approaches, along with specific cell autonomous and non-cell autonomous reagents), will be applied to prospective and non-prospective neural crest tissues before and after induction (specific aim 2);and finally we will characterize the interaction between Pax7 and Ubc9, and their role during neural crest development, (specific aim 3).