Sorting motifs are involved in the transport of diverse proteins. In the present study, we identified a hydrophobic peptide (WRPWRNFWWSIRVPWRRN) that was able to target enhanced green fluorescent protein- or DsRed2-enriched vesicular-like sub-compartments of the endoplasmic reticulum (ER). Analysis of mutation constructs revealed that the sequence WRPWRNFVVW was responsible for the ER targeting activity, and the arginine residue of the peptide is a critical determinant of ER localization. Results from co-immunoprecipitation, glutathione S-transferase pull-down, liquid chromatography tandem mass spectrometry, and western blotting analyses demonstrated that this motif could bind with the gamma 2-COP subcomplex of coat protein complex I (COPT), which is involved in the retrieval and transport of ER-resident proteins from the Golgi apparatus to the ER. Overall, we report a new hydrophobic peptide that possesses an arginine-based ER localization motif, which can help elucidate the mechanisms of ER sorting mediated by COPI. (C) 2016 Elsevier Inc. All rights reserved.