Embryonic determination is mediated in part by morphogenetic substances specifically localized in the cytoplasm of the egg. The vegetal region of eggs in general and frog eggs in particular is of special interest because two known determinants, those specifying the germ cell lineages and those inducing the dorsal axial structures, are known to reside in this region. Recent evidence suggests that mRNAs could function as determinants. This laboratory has found that a small group of mRNAs are localized to the vegetal pole of Xenoupus oocytes and has cloned one such RNA (Veg2). Veg2 RNA is specifically associated with the oocyte's cytoskeleton. In this proposal experiments are outlined to (1) determine when Veg2 RNA is expressed during Xenopus development and to characterize its protein product; (2) examine the pattern of inheritance of Veg2 RNA and its protein product during normal and perturbed embryogenesis; (3) establish the requirements for correct localization of Veg2 RNA to the vegetal pole; (4) explore the relationship of Veg2 RNA to the oocyte's cytoskeletion; (5) initiate studies designed to assay for the developmental function of Veg2 RNA; (6) determine if other vegetally localized mRNAs are associated with the cytoskeleton. The general strategy involves using the Veg2 cDNA clone in blot and in situ hybridization procedures to follow its developmental fate at the cellular level. Microinjection of Veg2 RNA into the animal pole of normal and perturbed eggs will yield information concerning the mechanisms of message localization. Antibodies will be raised to Veg2 protein produced in an expression vector system and used to characterize the Veg2 protin as to its biochemical nature and its possible function in development.
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