Weisblat 9723114 Glossiphoniid leeches such as Helobdella robusta, are useful for in depth analyses of developmental processes for two reasons: first, they are relatively simple animals with large, accessible embryos that are suited for combined cellular and molecular techniques. Second, since leeches belong to the phylum Annelida, an otherwise poorly studied group, the results obtained are useful for making interphyletic comparisons. Such comparisons are essential to distinguish general developmental processes from phylum-specific ones, and also to learn how evolutionary modification of developmental processes leads to the appearance of diverse animal types. For example, annelids and arthropods resemble each other (and differ from mollusks) in that they both have segmented body plans. In both annelids and arthropods, segmental tissues arise from longitudinally oriented, ventrally situated arrays of cells, termed the germ band (in arthropods such as the fruitfly Drosophila melanogaster) or germinal plate (in annelids). In leech, the germinal plate arises in strict head-to-tail (rostrocaudal) progression by repeated divisions of specialized embryonic stem cells called teloblasts. Homologs of several genes involved in rostrocaudal patterning of Drosophila seem to be expressed in homologous patterns during rostrocaudal patterning of leech. This suggests that the molecular genetic processes of differentiation and morphogenesis in later development will be quite similar in the two phyla. But the early Helobdella embryo is strikingly different from the early Drosophila embryo and strikingly similar to molluscan embryos. Thus, the question of how cell fates are determined in the early leech embryo is of considerable interest. The first part of this proposal contains experiments designed to further previous investigations of how distinct fates are assigned to particular cells in the developing embryo, with emphasis on the assignment of mesodermal and ectodermal fates to individually iden tified cells at fourth cleavage. The focus in this work is on characterizing the patterns of expression and development roles of leech homologs to evolutionarily conserved regulatory genes from Drosophila, such as nanos, dorsal, twist, snail, and numb. A second area of interest is the morphogenetic cell movements leading to germinal plate formation in Helobdella. Columns of segmental founder cells coalesce into the germinal plate having moved over most of the surface of the embryo; these movements are accompanied by the spreading of a squamous epithelium in a process called epiboly. The experiments in the second part of this proposal are designed to elucidate the cellular processes involved in these movements and to compare the mechanisms of epiboly in leech and fish. Specifically, we seek to test the hypothesis that the segmental founder cells and epithelium are "towed" vegetally by forces originating within the endodermal precursor cells; if so, we further seek to distinguish between possible modes of generating the towing movements, such as "cortical treadmilling" and "cortical contractile wave" processes.
