The transformation of a fertilized egg into a fully developed organism is a complex process, the molecular basis of which is just beginning to be revealed. Recent advances in recombinant DNA methodology, coupled with powerful genetic and embryological techniques, have led to the identification of a small number of regulatory genes which play pivotal roles in orchestrating the developmental program in vertebrate embryos. This proposal focuses on one of these genes called 1mx-1a. Molecular analysis of 1mx-1a indicates that it is a LIM- homeodomain transcription factor, and functions to regulate the synthesis of other , as yet undetermined, gene products. 1mx-1a is produced in discrete subsets of cells of the developing limb and the central nervous system during early stages of their formation, before any sign of overt differentiation has occurred. Based on its expression pattern in chick and mouse embryos, 1mx-1a is likely to play a primary regulatory role in the specification and coordination of cell fates in the limb and brain. In order to define the function of 1mx-1a in the development of limbs and of the mid-hindbrain region of the central nervous system, mice lacking this gene will be generated using embryonic stem cell technologies. The development of these mice will be studied using a variety of molecular and histological methods to define defects which arise from deletion of 1mx-1a. In addition, 1mx-1a will be expressed outside of its normal boundaries in chick embryos using recombinant avian viral vectors. The effect of this manipulation on the development of the chick mid-hindbrain region will be determined using molecular and anatomical markers of cell fate. Finally, the expression of 1mx-1a will be examined in mice with defects in limb and mid-hindbrain development to determine whether 1mx-1a might be involved in the generation of these lethal phenotypes. It is expected that these studies will greatly enrich our current understanding of vertebrate developmental mechanisms. Furthermore, the reagents generated as a result of this work will facilitate future endeavors aimed at defining specific molecular genetic processes which underlie the acquisition of shape and form during animal embryogenesis. Finally, these studies bear directly on human embryonic development since developmental mechanisms, including specific gene products, are well conserved among all vertebrates. Therefore, it is not unreasonable to assume that this work may lead to novel modalities aimed at the diagnosis and prevention of developmental anomalies of the limbs and brain.
