Protein trafficking events in the secretory and endosomal apparatus were evaluated in rat liver hepatocytes by EM immunogold cytochemistry. The cellular distribution of apolipoprotein E and other abundant hepatic secretory and endosomal proteins was quantitatively examined in ultrathin cryosections of liver hepatocytes. Under steady-state conditions, apoE was concentrated within the Golgi apparatus, along sinusoidal plasma membrane microvilli and within all components of the endosomal apparatus, as evaluated immunocytochemically and confirmed by quantitative immunoblotting of organelles isolated from liver homogenates. / Within the secretory pathway, the hepatic Golgi apparatus was a site of protein concentration as evaluated by the gold labeling density of another major secretory protein of liver hepatocytes, albumin, which was concentrated $ sim$10-fold in the Golgi apparatus relative to the ER. Sorting of this secretory protein within pre-Golgi compartments was not observed. Within the Golgi apparatus, apoE was concentrated within Golgi saccular distensions while being predominantly absent from flattened saccular components; apoB was similarly segregated within peripheral distensions. In contrast albumin, as well as two other monomeric proteins, transferrin (Tf) and the polymeric immunoglobulin receptor (pIg-R) were distributed homogeneously throughout Golgi stacks. In an attempt to assess a key prediction of the vesicular transport hypothesis, small 60-90 nm vesicles in the immediate vicinity of Golgi apparatus, postulated to mediate intersaccular transport were examined for their content of cargo secretory or plasma membrane proteins. Lack of immunoreactive apoE, apoB, albumin, Tf, or pIgR, within small vasicular profiles suggests limits to current models of vesicle-mediated intra-Golgi transport. / Along the endocytic pathway, at the cell surface, apoE and pIgR were dispersely distributed along the sinusoidal microvilli. Quantitative analysis of the immunolabeling distribution of these proteins did not reveal concentration within plasma membrane pits. These findings which were confirmed by observations of cell surface labeling of two other ligands, Tf and apoB, are consistent with receptors and ligands gaining access to the endocytic machinery likely without receptor/ligand preclustering or prolonged clustering events within plasma membrane pits. Intracellularly, apoE was concentrated within endocytic structures which were double-labeled for apoE and internalized HRP. Large endocytic vesicles closely juxtaposed to Golgi stacks also revealed a high content of apoE. Together the endosomal labeling distribution of apoE as well as morphological features of endosomal components are consistent with the maturation model for endosomes.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.29387 |
| Date | January 1995 |
| Creators | Dahan, Sophie |
| Contributors | Bergeron, John J. M. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Anatomy and Cell Biology.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001468241, proquestno: NN08090, Theses scanned by UMI/ProQuest. |
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