This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.COPI vesicles arise from Golgi cisternae and mediate the recycling of proteins from the Golgi back to the endoplasmic reticulum (ER), and the transport of Golgi resident proteins between cisternae. In vitro studies have produced evidence for two distinct types of COPI vesicles, but the in vivo sites of operation of these vesicles remain to be established. We have used a combination of electron tomography and immunolabeling techniques to examine Golgi stacks and associated vesicles in the cells of the scale-producing alga Scherffelia dubia and Arabidopsis preserved by high-pressure freezing/freeze-substitution methods. Five structurally distinct types of vesicles were distinguished. In Arabidopsis, COPI and COPII vesicle coat proteins as well as vesicle cargo molecules (mannosidase I and sialyltransferase-YFP) were identified by immunogold labeling. In both organisms the COPI-type vesicles were further characterized by a combination of six structural criteria: coat architecture, coat thickness, membrane structure, cargo staining, cisternal origin, and spatial distribution. Using this multi-parameter structural approach, we can distinguish two types of COPI vesicles, COPIa and COPIb. COPIa vesicles bud exclusively from cis cisternae and occupy the space between cis cisternae and ER export sites, whereas the COPIb vesicles bud exclusively from medial- and trans-Golgi cisternae and are confined to the space around these latter cisternae. We conclude that COPIa vesicle-mediated recycling to the ER occurs only from cis cisternae, that retrograde transport of Golgi resident proteins via COPIb vesicles is limited to medial and trans cisternae, and that diffusion of peri-Golgi vesicles is restricted.Donohoe, B.S., B.-H. Kang and L.A. Staehelin (2007) Identification and characterization of COPIa- and COPIb-type vesicle classes associated with plant and algal Golgi. Proc. Nat. Acad. Sci. USA 104:163-168.
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