Proteins traversing or resident in the secretory pathway of mammalian cells are found in the endoplasmic reticulum, Golgi apparatus, lysosomes, constitutive and regulated secretory vesicles, plasma membrane, and extracellular space. In this article, we review how proteins enter this pathway and move between compartments, how their proper folding and assembly are ensured by a quality-control system in the endoplasmic reticulum, and how they are modified as they move through the pathway. The mechanisms mediating protein localization in the endoplasmic reticulum, Golgi apparatus, and lysosomes are also discussed, as is the role of cytoskeletal proteins and molecular motors in intracellular trafficking.