BRAIN MORPHOGENESIS DEFECTS IN CEREBUM MUTANT ZEBRAFISH

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The period of gastrulation and neurulation is a particularly vulnerable one in vertebrate development, and genetic and environmental perturbations of these processes lead to a variety of congenital malformations in the human nervous system. Our understanding of normal and abnormal nervous system development has largely come from the study of animal model systems. My ultimate goal is to use a genetic approach in animal model systems to study fundamental processes in vertebrate neural development, and use my knowledge in the training of future physicians and medical researchers. In the present proposal, I outline an integrated program of research training, teaching, and independent research, designed to further my goals. I am pursuing postdoctoral training at the Carnegie Institution, an intensive basic research environment, in which I will learn a new animal model system and set of techniques. I will gain structured teaching experience, by lecturing in developmental biology and genetics courses that are team taught to the medical and graduate students at the Johns Hopkins University School of Medicine, and will also take advantage of opportunities offered at the Carnegie Institution to give seminars and journal clubs. In my proposed independent research project, I will examine aspects of early neural development in the teleost fish Danio rerio, the zebrafish. Easily accessible embryos and refined genetic techniques make the zebrafish an attractive system to study the genetic regulations underlying early development of the vertebrate nervous system. A new zebrafish mutant, cerebum (crb) has been isolated based on its dramatically reduced brain and eyes at 24 hours, and enlarged tail bud. I hypothesize that the primary defect in crb is in the convergent movement of cells during gastrulation; hence, cells destined for anterior neural and possibly mesodermal structures are found in the posterior region. I propose to characterize the crb mutation using embryologic, genetic, and molecular approaches. I will obtain new alleles of crb, and will use a combination of mapping and subtractive hybridization to isolate the crb gene. I will perform in situ hybridization for specific markers, to evaluate the patterning and development of the nervous system and dorsal mesoderm in crb mutants. Time-lapse analysis of cell behavior will complement the in situ hybridizations by yielding information on cell movements, proliferation, and lineage. Through cell transplants between crb and wild-type embryos I will determine the cell-autonomous nature of crb action. Finally I will examine epistatic interactions between crb and other mutations that effect cell movements in gastrulation by generating double mutants. This analysis should provide new information about the behavior of cells underlying the morphogenesis of the neural plate.