Regulated motion of glycoproteins revealed by direct visualization of a single cargo in the endoplasmic reticulum.
The quality of cargo proteins in the endoplasmic reticulum (ER) is affected by their motion during folding. To understand how the diffusion of secretory cargo proteins is regulated in the ER, we directly analyze the motion of a single cargo molecule using fluorescence imaging/fluctuation analyses. We find that the addition of two N-glycans onto the cargo dramatically alters their diffusion by transient binding to membrane components that are confined by hyperosmolarity. Via simultaneous observation of a single cargo and ER exit sites (ERESs), we could exclude ERESs as the binding sites. Remarkably, actin cytoskeleton was required for the transient binding. These results provide a molecular basis for hypertonicity-induced immobilization of cargo, which is dependent on glycosylation at multiple sites but not the completion of proper folding. We propose that diffusion of secretory glycoproteins in the ER lumen is controlled from the cytoplasm to reduce the chances of aggregation.
Nagaya
Hisao
H
Department of Cell Science, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima 960-1295, Japan.
Tamura
Taku
T
Higa-Nishiyama
Arisa
A
Ohashi
Koji
K
Takeuchi
Mayumi
M
Hashimoto
Hitoshi
H
Hatsuzawa
Kiyotaka
K
Kinjo
Masataka
M
Okada
Tatsuya
T
Wada
Ikuo
I
D008841:Actin Cytoskeleton / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure; D036681:Fluorescence Recovery After Photobleaching / Q000379:methods; D006023:Glycoproteins / Q000378:metabolism*; D006031:Glycosylation; D008164:Luminescent Proteins / Q000032:analysis; D008954:Models, Biological; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D009994:Osmolar Concentration; D017510:Protein Folding; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000032:analysis
D006023:Glycoproteins; D008164:Luminescent Proteins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins