ELECTRON TOMOGRAPHIC CHARACTERIZATION OF CIS-GOLGI CISTERNAL ASSEMBLY

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The Golgi of Scherffelia dubia has become a model for the study of intra-Golgi transport by cisternal maturation. This alga produces scales in its Golgi stacks. Scales are seen only inside the Golgi cisternae and not in the surrounding vesicles. This observation is a cornerstone of the cisternal maturation hypothesis. To further test the structural predictions of this hypothesis, we have produced 3-D Golgi reconstructions of high-pressure frozen and freeze-substituted cells at approximately 6 nm resolution using electron tomography. This analysis has allowed a detailed characterization and quantification of the structures involved in cis-cisterna assembly. Each transitional ER domain displays an average of nine approximately 70 nm COPII-type budding profiles. Approximately 30% of the COPII-type vesicles form tethered clusters of 3-5 vesicles in the space between the transitional ER and the cis-most Golgi cisterna. After attachment to the C2 cisterna, the tethered COPII-type vesicles appear to fuse to create cisternal initials that grow into a new cisterna by fusion with individual COPII-type vesicles. Putative docking/fusion complexes deform vesicles into a cone shape before they fuse with cisternal membranes. Two types of linker complexes connect growing cis-cisternae and one type connects medial- and trans-cisternae. Chlamydomonas and higher plant Golgi exhibit the same features, suggesting that they too traffic by cisternal maturation. Donohoe, BS, ZR Gergely, M. Gerl and LA Staehelin (2005) Electron tomographic characterization of cis-Golgi cisternal assembly and associated COPII vesicle budding and fusion events in Scherffelia dubia, Chlamydomonas reinhardtii, and higher plants. In review Mol.Biol. of Ce