The Drosophila gene cappuccino (capu) is related to the vertebrate limb deformity locus, and is required during Drosophila oogenesis to correctly localize developmental determinants within the oocyte. The microtubules in capu mutants are abnormally organized, resulting in premature microtubule-based cytoplasmic streaming and abnormal cell polarity. This suggests that capu either directly or indirectly regulates the cicrotubules. We now have a number of pieces of evidence indicating that CAPU functions dirctly in the actin cytoskeleton by binding to profilin, a small actin-binding protein. This proposal is aimed at further elucidating the eellular function of CAPU. We will isolate and analyze the phenotype of mutations that eliminate CAPU. We Will examine CAPU's distribution throughout development with epitope tagging and immunolocalization. To further define the physical interaction between CAPU and profilin and to obtain evidence that this interaction occurs in flies, we have four approaches. We will analyze additional double mutants between capu and chic, which encodes profilin, and between capu and other mutants that affect the actin cytoskeleton. We will map the domain of CAPU responsible for the profilin interaction. We will identiy or isolate mutants that specifically affect the interaction. These mutants will be moved into flies and we will attempt to alter the phenotype of these mutants by overexpressio of the second interactor. We will complete the molecular analysis of spir, a gene with a phenotype identical to capu and will isolate SPIR antibodies to determine the SPIR distribution in Wild-type and in spir, capu, and chic. To identify genes that might act with capu, we will screen for deficencies and then genes that exhibit second site non-complementation with capu.