Bulk movement

Abstract: The endoplasmic reticulum (ER) is the port of entry of the protein secretory pathway. Proteins destined for the cell wall, the vacuole or for the other compartments of the endomembrane system are first inserted into the ER and then transported to the Golgi complex en route to their final destinations. The ER is the compartment where newly-synthesized polypeptides fold, where many multimeric proteins assemble and where glycoproteins acquire their asparagine-linked glycans. The ER also provides a protein quality control function and proteins are usually retained in this compartment until they have acquired their correct conformation

Abstract: Many endoplasmic reticulum (ER) proteins maintain their residence by dynamic retrieval from downstream compartments of the secretory pathway. In previous work we compared the retrieval process mediated by the two signals, KKMP and KDEL, by appending them to the same neutral reporter protein, CD8, and found that the two signals determine a different steady-state localization of the reporter. CD8-K (the KDEL-bearing form) was restricted mainly to the ER, whereas CD8-E19 (the KKMP-bearing form) was distributed also to the intermediate compartment and Golgi complex. To investigate whether this different steady-state distribution reflects a difference in exit rates from the ER and/or in retrieval, we have now followed the first steps of export of the two constructs from the ER and their trafficking between ER and Golgi complex. Contrary to expectation, we find that CD8-K is efficiently recruited into transport vesicles, whereas CD8-E19 is not. Thus, the more restricted ER localization of CD8-K must be explained by a more efficient retrieval to the ER. Moreover, because most of ER resident CD8-K is not O-glycosylated but almost all CD8-E19 is, the results suggest that CD8-K is retrieved from the intermediate compartment, before reaching the Golgi, where O-glycosylation begins. These results illustrate how different retrieval signals determine different trafficking patterns and pose novel questions on the underlying molecular mechanisms.

Abstract: Information is carried around the nervous system by cells called neurons. The ability of neurons to communicate with each other relies on many proteins that are found on the surfaces of the cells. Like in all animal cells, surface proteins are made inside the cell in a compartment called the endoplasmic reticulum. During this process, one or several complex sugar molecules are usually added to newly made proteins. These sugar molecules are then modified as the proteins leave the endoplasmic reticulum and pass through another compartment called the Golgi apparatus on the way to the cell membrane. The precise number and structure of the sugar molecules attached to the protein define its glycosylation profile. Neurons receive information from other neurons at branch-like structures called dendrites, which trigger electrical signals that travel through the rest of the cell. To directly control how dendrites generate these signals, neurons make surface proteins locally in dendrites. However, while the endoplasmic reticulum is found all over the neuron, including in the dendrites, the Golgi apparatus is generally only present in the main cell body. It is not known how surface proteins are made in the dendrites or how the proteins’ glycosylation profiles are altered in the absence of a Golgi apparatus. Hanus et al. used microscopy and biochemical techniques to study the glycosylation profiles of surface proteins in rat neurons. The experiments revealed that immature glycosylation profiles are found on hundreds of different proteins that have been transported to the cell surface. This includes many proteins that are needed to transmit electrical signals between neurons. Next, Hanus et al. selectively blocked the modification of sugar molecules on proteins in the Golgi apparatus. This showed that dendrites are able to form and work properly even if surface proteins have primarily immature glycosylation profiles. Further experiments suggest that immaturely glycosylated proteins might travel to the surface of neurons using a different route that bypasses the Golgi apparatus. The next step will be to investigate exactly how these proteins are delivered to the surface and how they influence the way neurons behave.

Abstract: Glycosylphosphatidylinositol (GPI)-anchored proteins exit the ER in distinct vesicles from other secretory proteins, and this sorting event requires the Rab GTPase Ypt1p, tethering factors Uso1p, and the conserved oligomeric Golgi complex. Here we show that proper sorting depended on the vSNAREs, Bos1p, Bet1p, and Sec22p. However, the t-SNARE Sed5p was not required for protein sorting upon ER exit. Moreover, the sorting defect observed in vitro with bos1-1 extracts was also observed in vivo and was visualized by EM. Finally, transport and maturation of the GPI-anchored protein Gas1p was specifically affected in a bos1-1 mutant at semirestrictive temperature. Therefore, we propose that v-SNAREs are part of the cargo protein sorting machinery upon exit from the ER and that a correct sorting process is necessary for proper maturation of GPI-anchored proteins.