Proteomic Profiling of Mammalian COPII and COPI Vesicles

Intracellular transport and homeostasis of the endomembrane system in eukaryotic cells depend on the formation and fusion of vesicular carriers. Coat protein complex (COP) II vesicles export newly synthesized secretory proteins from the endoplasmic reticulum (ER), whereas COPI vesicles facilitate traffic from the Golgi to the ER and intra-Golgi transport. Mammalian cells express various isoforms of COPII and COPI coat proteins. To investigate the roles of coat protein paralogs, we have combined in vitro vesicle reconstitution from semi-intact cells with SILAC-based mass spectrometric analysis. Here, we describe the core proteomes of mammalian COPII and COPI vesicles. Whereas the compositions of COPII vesicles reconstituted with various isoforms of the cargo-binding subunit Sec24 differ depending on the paralog used, all of the isoforms of the COPI coat produce COPI-coated vesicles with strikingly similar protein compositions. [Display omitted] •SILAC-based proteomics reveals the proteomes of mammalian COPI and COPII vesicles•ERGIC1, a putative cycling cargo adaptor, is an Sec24C/D-dependent cargo protein•CNIH4, an adaptor that controls GPCR exit from the ER, is an Sec24A-dependent client•COPI vesicles produced with different γ/ζ-COP and Arf paralogs have similar proteomes Intracellular transport between the ER and the Golgi is archived by COPI and COPII vesicles. Adolf et al. used an in vitro reconstitution assay in combination with SILAC-based mass spectrometric analysis to reveal the core proteome of these transport vesicles and to define the role of coat protein isoforms.