| Subcellular asymmetry, or cell polarity, is essential to eukaryotic cells for diverse biological processes such as cell division, differentiation, and morphogenesis. The budding yeast, Saccharomyces cerevisiae, depends on cell polarity for vegetative growth, cell division and mating. Spa2p, Pea2p, Bud6p/Aip3p, and Bni1p are known to be involved in regulating cell polarity during budding and mating. These proteins localize to sites of polarized growth, share polarity-related mutant phenotypes, and associate with each other by various criteria. However, the molecular functions of these proteins are still unknown. In order to gain insight into the roles of this set of proteins in polarity development, both genetic and biochemical approaches were used. First, a co-immunoprecipitation method was used to identify proteins that associate with Spa2p. The yeast myosin proteins, Myo1p and Myo2p, were identified by this method, and the significance of their interaction with Spa2p was further characterized. We uncovered a genetic interaction between Spa2p and Myo1p, suggesting that Spa2p may contribute to the cell-separation and cytokinesis functions of Myo1p. We also discovered that Spa2p co-fractionated with Myo2p and associated with actin in an ATP-sensitive manner, presumably via its interaction with Myo2p. Furthermore, Spa2p and Myo2p depended on each other for proper localization to sites of polarized growth. We propose that Myo2p skin contained this consensus sequence. To confirm these database predictions of biological substrates, potential substrates were digested with pure cercarial elastase in vitro and skin sections incubated with pure cercarial elastase.;Cercarial elastase has previously been shown to be immunogenic in humans. In collaboration with Dr. Reda Ramzy (Ain Shams University, Cairo, Egypt) an ELISA was developed using purified native cercarial elastase as antigen. Results of testing the ELISA in endemic villages indicated there is a positive correlation between disease incidence and cercarial elastase antibodies. This suggests that the ELISA will be a useful epidemiological tool to identify exposure to schistosome larvae. |
