Maturation of dense core granules in wild type and mutant Tetrahymena thermophila. |
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| Authors: | A P Turkewitz L Madeddu R B Kelly |
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| Affiliation: | Department of Biochemistry and Biophysics, University of California, San Francisco 94143. |
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| Abstract: | Tetrahymena thermophila, a ciliated protozoan, has a well-developed pathway of regulated secretion from dense core granules called mucocysts. Since exocytosis-defective mutants are available, steps in the biogenesis of dense core granules and their fusion with the plasma membrane may be resolved genetically. To describe the steps in biochemical terms, we have generated antisera against mucocyst content proteins. One antiserum is directed against a calcium binding protein, p40, that is released on stimulation of exocytosis. p40 is shown to associate with an insoluble matrix in mature mucocysts. In addition, the antiserum recognizes a larger protein, p60, that is soluble, is not found in mature mucocysts and is not released on stimulation. Pulse-chase experiments support a precursor-product relationship between p60 and p40. Using these proteins as markers, two mutant Tetrahymena strains defective in exocytosis have been shown to accumulate the putative precursor p60 in organelles that can be distinguished from one another and from wild type mucocysts on the basis of density. The kinetics of appearance of insoluble p40 and the mutant phenotypes suggest a model of mucocyst maturation in which sorting precedes matrix condensation. |
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