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Title: Retention and targeting in the Golgi apparatus
Author: Hui, Hu
Awarding Body: University of London
Current Institution: University College London (University of London)
Date of Award: 1997
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A matrix that binds medial Golgi enzymes can be isolated as a detergent insoluble complex. Some components of this complex were identified as cytoskeletal proteins, e.g. cytokeratins and actin. To further analyse this matrix, Golgi membranes were extracted in detergent and salt followed by dialysis and fractionation on a linear sucrose density gradient. An oligomer was identified with an apparent molecular weight of 2x106D as assessed by size-exclusion chromatography. It contained two medial Golgi enzymes, α-1,3-1,6-mannosidase II (Mann II) and β-1,2-N-acetylglucosaminyltransfer-ase I (NAGT I), along with various other proteins. Some of the components of the oligomer were identified as being proteins homologous to the lectins, ERGIC53/p58 and VIP36, and members of the p24 family of putative cargo receptors. The isolation of the oligomer and the identification of some of its components represent a step closer to the elucidation of the mechanisms of enzyme retention and the structural maintenance of the Golgi apparatus. The early Golgi t-SNARE, syntaxin 5, is thought to provide targeting specificity for both COPI and COPII vesicles originating from the endoplasmic reticulum (ER) and COPI vesicles on the retrograde pathway. Two forms of syntaxin 5 were shown to be generated from the same mRNA by alternative initiation of translation. The short form (35kD) corresponds to the published sequence (Bennett et al. (1993), Cell, 74, 863-73). The longer form (42kD) is novel, with an N-terminal, cytoplasmic extension, containing a predicted type II ER targeting signal. When grafted onto a reporter molecule, this signal localised the construct to the ER as assessed by immuno-fluorescence microscopy. This signal could function to retrieve syntaxin 5 from later Golgi compartments but several lines of evidence, including the absence of this longer form from the yeast homologue, Sed5p, point to a function unique to higher organisms.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available