Figure 7.
Schematic representation for the localization and function of the COPII heterocomplex at the ER–ERES boundary: Support for direct ERES to Golgi transport. (A) The localization and function of the COPII heterocomplex at the ER–ERES boundary. Main panel: COPII dynamically binds and establishes domains of collar-like elongated membranes that comprise a stable ER–ERES boundary. Cargo accumulates in COPII-free ERES membranes by passage through the COPII coated neck. Fission ensues to form Golgi bound COPII-deficient carriers. Top right: Hydrophobic mismatching between transmembrane domains of cargo proteins and surrounding lipids is essential for cargo sorting: Schematic representation demonstrating how bilayer thickness gradient from thinner ER to thicker ERES membranes drives cargo into ERES. Alleviation of hydrophobic mismatching of cargo transmembrane domains facilitates their concentration in ERES membranes by preventing their diffusion through the COPII neck back to the ER. Bottom right: A detailed model of the COPII cargo sorting machinery at the ER–ERES boundary. COPII is recruited exclusively at the ER adjacent to the COPII neck by Sec12 and Sar1-GTP and binds cargo via the Sec24 subunit. The COPII-cargo complex is driven toward ERES potentially by the abovementioned hydrophobic mismatch interactions. COPII coat disassembly initiated by Sar1-GTP hydrolysis releases the cargo at the distal ERES end of the COPII-coated neck. (B) Golgi-associated ERESs in living intact cells. The intracellular distribution of COPII subunit Sec24C-mCherry (red) and Rab1b-YFP were used here as a Golgi marker. A confocal image of the Huh7 cell coexpressing the Rab1b-YFP (left, and green in merged image) and the COPII subunit Sec24C-mCherry (center, and red in merged image). Golgi apparatus marked by G. The frame showing the Golgi apparatus is enlarged threefold below. Scale bar = 5 µm. (C) Stable Golgi-associated ERES during cargo accumulation in Golgi apparatus. Time-lapse analysis of living cells coexpressing Sec24C-mCherry (red) and VSVG-YFP (green), showing an accumulation of cargo VSVG-YFP in Golgi membranes decorated with COPII-labeled domains. The area in white rectangles is magnified 3.3-fold on the bottom. Scale bars in top and bottom panels are 10 and 5 µm, respectively. (D) Golgi-associated ERESs disappear upon BFA-induced Golgi membranes blink-out. Representative images from a time-lapse sequence taken at 3-s intervals after addition of 5 µg/ml BFA to living cells coexpressing GalT-YFP (green, bottom) and Sec24C-mCherry (red and inverted, middle). Times are counted. Scale bar = 5 µm.

Schematic representation for the localization and function of the COPII heterocomplex at the ER–ERES boundary: Support for direct ERES to Golgi transport. (A) The localization and function of the COPII heterocomplex at the ER–ERES boundary. Main panel: COPII dynamically binds and establishes domains of collar-like elongated membranes that comprise a stable ER–ERES boundary. Cargo accumulates in COPII-free ERES membranes by passage through the COPII coated neck. Fission ensues to form Golgi bound COPII-deficient carriers. Top right: Hydrophobic mismatching between transmembrane domains of cargo proteins and surrounding lipids is essential for cargo sorting: Schematic representation demonstrating how bilayer thickness gradient from thinner ER to thicker ERES membranes drives cargo into ERES. Alleviation of hydrophobic mismatching of cargo transmembrane domains facilitates their concentration in ERES membranes by preventing their diffusion through the COPII neck back to the ER. Bottom right: A detailed model of the COPII cargo sorting machinery at the ER–ERES boundary. COPII is recruited exclusively at the ER adjacent to the COPII neck by Sec12 and Sar1-GTP and binds cargo via the Sec24 subunit. The COPII-cargo complex is driven toward ERES potentially by the abovementioned hydrophobic mismatch interactions. COPII coat disassembly initiated by Sar1-GTP hydrolysis releases the cargo at the distal ERES end of the COPII-coated neck. (B) Golgi-associated ERESs in living intact cells. The intracellular distribution of COPII subunit Sec24C-mCherry (red) and Rab1b-YFP were used here as a Golgi marker. A confocal image of the Huh7 cell coexpressing the Rab1b-YFP (left, and green in merged image) and the COPII subunit Sec24C-mCherry (center, and red in merged image). Golgi apparatus marked by G. The frame showing the Golgi apparatus is enlarged threefold below. Scale bar = 5 µm. (C) Stable Golgi-associated ERES during cargo accumulation in Golgi apparatus. Time-lapse analysis of living cells coexpressing Sec24C-mCherry (red) and VSVG-YFP (green), showing an accumulation of cargo VSVG-YFP in Golgi membranes decorated with COPII-labeled domains. The area in white rectangles is magnified 3.3-fold on the bottom. Scale bars in top and bottom panels are 10 and 5 µm, respectively. (D) Golgi-associated ERESs disappear upon BFA-induced Golgi membranes blink-out. Representative images from a time-lapse sequence taken at 3-s intervals after addition of 5 µg/ml BFA to living cells coexpressing GalT-YFP (green, bottom) and Sec24C-mCherry (red and inverted, middle). Times are counted. Scale bar = 5 µm.

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