Conserved transport signals for exiting the endoplasmic reticulum in COPII-coated vesicles

As COPII vesicles bud from the endoplasmic reticulum (ER) they incorporate cargo and SNARE proteins. In the yeast COPII transport system three distinct sites on the COPII Sec23/24 sub-complex encode determinants for cargo and SNARE selectivity. In mammals, the level of complexity of cargo packaging increases, as there are multiple Sec23/24 isoforms. Our studies focus on a detailed understanding of the molecular determinants for SNARE and cargo export from the mammalian ER by the COPII vesicular coat complex. The results show that the SNARE Bet1 and VSV-G are selected by the Sec24A and Sec24B subunits at the conserved B-site, a binding pocket on the equatorial surface of Sec24. The SNAREs Syntaxin 5 and Membrin contain a novel 'IxM' signal that binds in an extended conformation to a conserved groove on the periphery of Sec24C and Sec24D. COPII vesicles budded with Sec24C and D isoforms package Syntaxin 5 and Membrin; however, Sec24C and D isoforms mutant at the IxM binding site do not package Syntaxin 5 and Membrin, validating the IxM sequence as a transport signal for exiting the ER.; The SNARE Sec22 has a bipartite signal that binds Sec23/24A and Sec23/24B and specifies its ER exit as an unassembled SNARE. The 200 kDa crystal structure of Sec22 bound to Sec23/24 reveals that the transport signal is a folded epitope rather than a conventional short peptide sequence. The SNARE motif folds against the N-terminal longin domain, and this closed form of Sec22 binds at the Sec23/24 interface. Thus, COPII recognizes unassembled Sec22 via a folded epitope, whereas Sec22 assembly into SNARE complexes would mask the SNARE motif. Our studies define the cargo selectivity and isoform specificity of the mammalian COPII vesicular coat complex.