Greater than Craniofacial malformations constitute one of the major groups of congenital defects in newborn human infants. The formation of the anterior aspect of the vertebrate embryo is a complex process for which we have few molecular details, especially regarding the contribution of the neural crest (NC). The NC is a transient structure composed of cells that migrate extensively throughout the embryo and contribute to a number of organs. The control of crest cell morphogenesis is critical to the embryo, in as much as abnormal NC migration or proliferation results in a number of serious human disorders ranging from cleft palate, to frontonasal dysplasia. The cranial malformations observed in children exposed in utero to 13-cis retinoic acid (RA) or excess alcohol are a consequence of the drugs' effect on some aspect of crest cell morphogenesis. The long term goal of the lab is to define the factors controlling normal cranial facial development and to elucidate the mechanism(s) by which retinoic acid and ethanol affect these processes. Our initial efforts focused on the NC of the mouse where we established CRABPI (cellular retinoic acid binding protein I) as a marker for a subpopulation of crest cells that contributed to many of the anterior structures of the embryo. The data we obtained indicated that these NC cells underwent dynamic movements during embryogenesis. Unfortunately, our studies have been limited by the inaccessibility of neural crest in the living embryo which is a consequence of the in utero development of mice. We have now refocused our efforts on zebrafish as a model system to study the role of the NC in craniofacial development. Since the embryos are transparent, the movement of individual cells can be visualized in real time. Like humans, the formation of the anterior structures of the zebrafish embryo is sensitive to teratogens like RA.
The specific aims of this proposal are: 1) clone and characterize the expression of zebrafish AP-2; and 2) to use the regulatory elements of AP-2 to generate transgenic lines of fish that express a fluorescent protein (GFP) in specific NC cell populations. This will allow us to follow the movement of AP-2+ cells in the living embryo. This last Aim represents a new direction for our laboratory. While we have extensive expertise in transgenesis in mice, the generation of transgenic fish will require the acquisition of new technologies for our group. Since our success in obtaining an R01 is dependent upon our having the transgenic fish, the funds from an NIDR Small Grant award will allow us to gain the required expertise, as well as establish and characterize the engineered lines of zebrafish.
