Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.643334
Title: Studies of Golgi organization and protein secretion in yeasts
Author: Colley, Alan D.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1995
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Abstract:
The PMR1 gene of Saccharomyces cerevisiae is predicted to encode a P-type Ca2+ ATPase (Rudolph et al., 1989). This protein has been reported as localizing to a novel Golgi-like organelle (Antebi and Fink, 1992). Consistent with Pmr1p's proposed Golgi distribution is the fact that the pleiotropic phenotype of null mutants results in defects in various Golgi processes (e.g. outer chain glycosylation, proteolytic processing, vacuolar sorting). These defects are reversible by addition of Ca2+ to the extracellular medium, supporting the proposed function as a Ca2+ pump (Rudolph et al., 1989; Antebi and Fink, 1992). Work described in this thesis was carried out to investigate the nature of the pmr1 phenotype and to further characterize the Pmr1p containing organelle(s). It is demonstrated that the pmr1 phenotype is not due to a complete bypass of the Golgi (as proposed by Rudolph et al., 1989) since Kex2p processing of a secreted protein is detected in a pmr1 mutant. There does however appear to be a change in Golgi organization in pmr1 mutants. When the organelles containing Kex2p were isolated from pmr1 and PMR1 strains the profiles of other marker enzymes recovered was significantly altered. In particular the enzyme GDPase, an early Golgi marker, colocalizes with Kex2p, a trans-Golgi network marker, in pmr1 strains. Furthermore DPAP A (Ste13p) no longer colocalizes with Kex2p in pmr1 strains. This evidence suggests that protein targeting and/or retention is altered in pmr1 strains as a result of aberrant Ca2+ homeostasis. Another possibility is that some gross reorganization of Golgi structure has occurred. These changes in enzyme localization were not reversible by addition of Ca2+ to the growth media. Pmr1p was tagged with protein A to allow isolation of organelles containing the fusion protein with IgG-Sepharose. However, after characterization of the recovered material it became clear that the fusion protein had localized to the vacuole and not the Golgi. This is probably due to the tag interfering with proper Golgi retention. This result is consistent with the vacuolar default model for membrane proteins in yeast (Roberts et al., 1992; Nothwher et al., 1993).
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.643334  DOI: Not available
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