EXOGENOUS RECEPTOR PROBES OF SIGNALLING IN EMBRYOGENESIS

Summary

Principal Investigator: WILLIAM BUSA
Abstract: Both classical and modern studies point toward a central role for cell-cell signalling during early pattern formation events in the frog embryo, the preeminent experimental model of vertebrate development. Such signalling may involve secreted growth factor- like molecules and their plasma membrane receptors, a suggestion based on demonstrations that certain "classical" growth factors have close homologs in the maternal mRNA pool of the embryo and can induce isolated ectoderm to form mesodermal cell types. Our long-term interest is the role of signal transduction (receptor- mediated "second messenger" production) in the early signalling events which establish the body plan of the vertebrate embryo. This essentially biochemical step in the pattern formation process, lying as it does between two molecular genetic steps (on the one hand, the localization of determinants such as mRNAs in cytoplasm destined to compose "transmitting" cells, and on the other, the modification of gene expression which is the outcome of induction) has received little serious study, yet it is at precisely this chemical step where, we may suppose, many human teratogens (e.g., lithium, retinoic acid, etc.) are likely to exert their developmental effects. Unfortunately, familiar crude approaches for manipulating signalling systems -- microinjecting putative second messengers, treating with pharmacological inhibitors or activators of enzymes involved in signal transduction, and so forth -- have only rarely proven fruitful in studies of early development in intact embryos, where problems of long-term nonspecific toxicity, precise timing of stimulation (and its termination), and spatially localized delivery are routinely encountered. We have recently demonstrated the feasibility of a unique new approach to this issue: by injecting the frog zygote or early embryo with mRNA encoding an exogenous hormone receptor which is rapidly and functionally expressed in the blastula, we can install molecular "switches" in the embryo and use them to specifically stimulate or inhibit various second messenger systems (singly or in combinations) with a high degree of both spatial and temporal resolution. This novel approach provides us an unprecedented opportunity to understand the roles of these signalling events in early pattern formation, as demonstrated by our initial finding of profound effects on dorso-anterior differentiation in frog embryos expressing exogenous serotonin receptors linked to polyphosphoinositide hydrolysis and intracellular calcium mobilization.
Funding Period: 1990-01-03 - 1993-12-31
more information: NIH RePORT