Abstract
SummaryEndoplasmic reticulum (ER)-to-Golgi anterograde transport is driven by COPII vesicles mainly composed of a Sec23/Sec24 inner shell and a Sec13/Sec31 outer cage. How COPII vesicles are tethered to the Golgi is not completely understood. We demonstrated here that PAQR3 can facilitate tethering of COPII vesicles to the Golgi. Proximity labeling using PAQR3 fused with APEX2 identified that many proteins involved in intracellular transport are in close proximity to PAQR3. ER-to-Golgi trafficking of N-acetylgalactosaminyltransferase-2 on removal of brefeldin A is delayed by PAQR3 deletion. RUSH assay also revealed that ER-to-Golgi trafficking is affected by PAQR3. The N-terminal end of PAQR3 can interact with the WD domains of Sec13 and Sec31A. PAQR3 enhances Golgi localization of Sec13 and Sec31A. Furthermore, PAQR3 is localized in the ERGIC and cis-Golgi structures, the acceptor sites for COPII vesicles. Taken together, our study uncovers a role for PAQR3 as a player in regulating ER-to-Golgi transport of COPII vesicles.
Highlights
Cellular homeostasis relies on the coordinated interactions of proteins and the correct localization of those proteins to specific subcellular compartments
As reported by previous studies (Feng et al, 2007; Luo et al, 2008), ascorbate peroxidase 2 gene (APEX2)-PAQR3 was mainly localized in the Golgi apparatus (Figure 1B)
We characterized the enzymatic activity of APEX2-PAQR3 fusion protein
Summary
Cellular homeostasis relies on the coordinated interactions of proteins and the correct localization of those proteins to specific subcellular compartments. Mapping these network and subcellular proteome associated with a protein is critical for revealing its biological functions. Proximity labeling is one of the most effective approaches that can obtain spatially resolved proteomic maps of specific proteins and compartments within living cells (Hung et al, 2016). Proximity labeling relies on a engineered ascorbate peroxidase (APEX), which is capable of generating free biotinyl radicals to enable a rapid and spatially restricted labeling of proteins in the vicinity of the enzyme (Bersuker et al, 2018; Kim and Roux, 2016; Mick et al, 2015). The irreversible conjugation of biotin catalyzed by APEX2 enables the capture of labeled proteins for proteomic analysis
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