Dr. Robinson proposes to study the development and expression of polarity in two systems: the zygotes of the fucoid alga, Pelvetia, and the early somatic embryos of carrot. He will focus on the ionic and electrical aspects of polarity, and the interactions of ions with the cytoskeleton. In the case of Pelvetia, he will use the fluorescent calcium indicator fura 2 to determine the spatial distribution of calcium ions in the zygote in order to resolve the controversy over the role of calcium gradients in polar axis development. Quantitative fluorescence studies, utilizing a video image processing system, will be carried out to determine the temporal relationships between calcium ion gradients, actin microfilament asymmetries, microtubule asymmetries and rhizoid formation. The causal relationships of these will also be explored. Dr. Robinson's results indicate that polarity in carrot somatic embryos is also associated with ionic currents and that protons are one component of the current. He will continue his studies to identify the current-carrying ions and the relationship of the ionic current to embryonic development and polar auxin transport. He will use antibodies to the plasma membrane H-ATPase to localize this protein at both the cellular and tissue level. He will also apply electrical fields to carrot protoplasts and determine the effects on the timing and plane of first cell division and the stimulation of embryogenesis. He will determine the distribution of the H-ATPase of protoplast plasma membranes in response to applied electrical fields. %%% There is evidence on both sides of the issue of whether endogenous electrical currents are important for the establishment of polarity in developing plant embryos. This establishment of polarity is the first event in formation of an asymmetric pattern in a previously symmetrical fertilized egg resulting ultimately in the asymmetric adult organism. This study it to attempt to clarify the involvement of these currents in this early patterning event.
