title: Organelle proteomics reveals cargo maturation mechanisms associated with Golgi-like encystation vesicles in the early-diverged protozoan Giardia lamblia.
abstract: During encystation Giardia trophozoites secrete a fibrillar extracellular matrix of glycans and cyst wall proteins on the cell surface. The cyst wall material is accumulated in encystation-specific vesicles (ESVs), specialized Golgi-like compartments generated de novo, after export from the endoplasmic reticulum (ER) and before secretion. These large post-ER vesicles neither have the morphological characteristics of Golgi cisternae nor sorting functions, but may represent an evolutionary early form of the Golgi-like maturation compartment. Because little is known about the genesis and maturation of ESVs, we used a limited proteomics approach to discover novel proteins that are specific for developing ESVs or associated peripherally with these organelles. Unexpectedly, we identified cytoplasmic and luminal factors of the ER quality control system on two-dimensional electrophoresis gels, i.e. several proteasome subunits and HSP70-BiP. We show that BiP is exported to ESVs and retrieved via its C-terminal KDEL signal from ESVs. In contrast, cytoplasmic proteasome complexes undergo a developmentally regulated re-localization to ESVs during encystation. This suggests that maturation of bulk exported cyst wall material in the Golgi-like ESVs involves both continuous activity of ER-associated quality control mechanisms and retrograde Golgi to ER transport.
authors.affiliation: Institute of Parasitology, University of Zürich, Winterthurerstrasse 266a, CH-8057 Zürich, Switzerland.
authors.forename: Sasa
authors.initials: S
authors.lastname: Stefanic
authors.forename: Daniel
authors.initials: D
authors.lastname: Palm
authors.forename: Staffan G
authors.initials: SG
authors.lastname: Svärd
authors.forename: Adrian B
authors.initials: AB
authors.lastname: Hehl
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002499:Centrifugation, Density Gradient; D003593:Cytoplasm / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D004229:Dithiothreitol / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D004721:Endoplasmic Reticulum / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D005819:Genetic Markers; D005822:Genetic Vectors; D016829:Giardia lamblia; D006056:Golgi Apparatus / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D013058:Mass Spectrometry; D008853:Microscopy; D008861:Microsomes / Q000378:metabolism; D008954:Models, Biological; D018832:Molecular Chaperones / Q000378:metabolism; D009154:Mutation; D009696:Nucleic Acids / Q000737:chemistry; D016366:Open Reading Frames; D011134:Polysaccharides / Q000737:chemistry; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D017434:Protein Structure, Tertiary; D040901:Proteomics / Q000379:methods*; D011994:Recombinant Proteins / Q000737:chemistry; D013347:Subcellular Fractions; D013395:Sucrose / Q000494:pharmacology; D013997:Time Factors; D019076:Transgenes
chemical_list: D004220:Disulfides; D000091342:Endoplasmic Reticulum Chaperone BiP; D005819:Genetic Markers; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D009696:Nucleic Acids; D011134:Polysaccharides; D011994:Recombinant Proteins; D013395:Sucrose; D046988:Proteasome Endopeptidase Complex; D004229:Dithiothreitol