9506094 Kropf The research described in this proposal will investigate the cytoskeletal and ionic bases of polarity in zygotes of the brown alga Pelvetia. The premise in this proposal is that the acquisition of polarity is a highly regulated reorganization of the cell which will be best understood by investigating relationships between key regulatory processes. Dr. Kropf will study cytoskeletal control of polarity, and then investigate the relationship between the cytoskeleton and cytosolic regulatory ions. Preliminary evidence indicates that cortical F-actin localizes in a patch soon after fertilization, and expands into a ring that portends the future rhizoid site. Dr. Kropf hypothesizes that sperm entry induces local F-actin polymerization that then serves as a determinant of polarity, specifically marking the future rhizoid site. This hypothesis will be examined using cryofixation, confocal microscopy and TEM. Prior to mitosis, the nucleus rotates 90 degrees bringing centrosomes into an axial alignment. This nuclear rotation determines the axial orientation of the spindle and transverse alignment of the first division plane. Microtubules are thought to be primarily responsible for nuclear rotation, and their dynamic behavior during rotation will be investigated in real time by injecting zygotes with fluorescently tagged tubulin. Centrosome assembly and function will also be investigated. Finally the hypothesis that the assembly and function of cytoskeletal elements are regulated by local ion activities, specifically H+ and CA+2 activities, will be investigated. ***
