Using STED super-resolution microscopy, researchers at the University of British Columbia have shed new light on the compartmentalization in peripheral ER tubules. Using the lumenal ER reporter ER monomeric oxidizing environment-optimized green fluorescent protein (ERmoxGFP), the team led by Dr. Ivan Nabi performed high-speed (40 ms/frame) live-cell STED imaging and characterized the behavior of the ER-shaping proteins. The results show that RTN and CLIMP-63 regulate the localization and dynamics of nanodomains along the tubules in very distinct ways. RTN segregates away from and restricts lumenal blob length, while CLIMP-63 associates with and increases lumenal blob length. ERmoxGFP defines local lumenal filling of nanodomains that are segregated from membrane-associated ER proteins along peripheral ER tubules. RTN and CLIMP-63 constitute key players for establishing ER nanodomain heterogeneity, interaction with ER proteins, and dynamics in ER tubules.
New Light Shed on the Nanodomain Organization of the Endoplasmic Reticulum (ER)
Information revealed with super-resolution STED nanoscopy
The endoplasmic reticulum (ER) is a continuous membrane organelle in charge of protein synthesis, lipid synthesis and detoxification. The ER structure is described in terms of smooth peripheral tubules and rough ER sheets. The family of proteins responsible for maintaining the sheet or tubule architecture include the cytoskeleton-linking membrane protein 63 (CLIMP-63) and reticulon (RTN), respectively. A detailed map of the nanodomain distribution remains challenging due to the size of the structures and their highly dynamic nature. To give an idea, the ER sheet thickness and ER tubule diameter are in the 30-100 nm range, well below the diffraction limit of confocal microscopy.