Identification of ERGIC-53 as an intracellular transport receptor of α1-antitrypsin

Abstract

Secretory proteins are exported from the endoplasmic reticulum (ER) by bulk flow and/or receptor-mediated transport. Our understanding of this process is limited because of the low number of identified transport receptors and cognate cargo proteins. In mammalian cells, the lectin ER Golgi intermediate compartment 53-kD protein (ERGIC-53) represents the best characterized cargo receptor. It assists ER export of a subset of glycoproteins including coagulation factors V and VIII and cathepsin C and Z. Here, we report a novel screening strategy to identify protein interactions in the lumen of the secretory pathway using a yellow fluorescent protein–based protein fragment complementation assay. By screening a human liver complementary DNA library, we identify α1-antitrypsin (α1-AT) as previously unrecognized cargo of ERGIC-53 and show that cargo capture is carbohydrate- and conformation-dependent. ERGIC-53 knockdown and knockout cells display a specific secretion defect of α1-AT that is corrected by reintroducing ERGIC-53. The results reveal ERGIC-53 to be an intracellular transport receptor of α1-AT and provide direct evidence for active receptor-mediated ER export of a soluble secretory protein in higher eukaryotes.

SNX 18 regulates ATG 9A trafficking from recycling endosomes by recruiting Dynamin‐2

“…To identify the membranes where the SNX18‐Dynamin‐2 interaction takes place, we took advantage of the split‐YFP bimolecular fluorescence complementation assay 35. In line with the IP results (Fig 3C), many YFP puncta were detected in cells expressing YFP1‐Dynamin‐2 and YFP2‐SNX18 (or YFP1‐SNX18 and YFP2‐Dynamin‐2) with only a few YFP spots seen in cells expressing the SNX18 W38K mutant (Fig 3F).…”

Crystal structure of the LMAN1‐CRD/MCFD2 transport receptor complex provides insight into combined deficiency of factor V and factor VIII

“…Many proteins have been shown to act as cargo for LMAN1 besides FV and FVIII e.g. cathepsin Z and cathepsin C [8,10], a1-antitrypsin [11], and sulfatase modifying factor 1 [12].…”

Inherited hematological disorders due to defects in coat protein (COP)II complex

“…Accordingly, cathepsin C, cathepsin Z and 1-antitrypsin have also been reported as potential cargo proteins dependent on LMAN1 for efficient secretion [73,74]. However, LMAN1-deficient mice, with reduced FV and FVIII plasma levels to 50% of wild-type mice, show no differences to wild type mice in the levels of cathepsin C and cathepsin Z in liver lysates or a1-antitrypsin levels in plasma [75].…”