Cargo receptor-assisted endoplasmic reticulum export of pathogenic α1-antitrypsin polymers

Abstract

Circulating polymers of α1-antitrypsin (α1AT) are neutrophil chemo-attractants and contribute to inflammation, yet cellular factors affecting their secretion remain obscure. We report on a genome-wide CRISPR-Cas9 screen for genes affecting trafficking of polymerogenic α1AT^H334D. A CRISPR enrichment approach based on recovery of single guide RNA (sgRNA) sequences from phenotypically selected fixed cells reveals that cells with high-polymer content are enriched in sgRNAs targeting genes involved in "cargo loading into COPII-coated vesicles," where "COPII" is coat protein II, including the cargo receptors lectin mannose binding1 (LMAN1) and surfeit protein locus 4 (SURF4). LMAN1- and SURF4-disrupted cells display a secretion defect extending beyond α1AT monomers to polymers. Polymer secretion is especially dependent on SURF4 and correlates with a SURF4-α1AT^H334D physical interaction and with their co-localization at the endoplasmic reticulum (ER). These findings indicate that ER cargo receptors co-ordinate progression of α1AT out of the ER and modulate the accumulation of polymeric α1AT not only by controlling the concentration of precursor monomers but also by promoting secretion of polymers.

Keywords: CHO CRISPR-Cas9 library; CHO cells; ERGIC-53; LMAN1; SURF4; cargo receptors; endoplasmic reticulum; genome-wide CRISPR-Cas9 screen; polymer trafficking; α1-antitrypsin.