Exploring the consequences of redirecting an exocytic Rab onto endocytic vesicles

Bidirectional vesicular traffic links compartments along the exocytic and endocytic pathways. Rab GTPases have been implicated in specifying the direction of vesicular transport. To explore this possibility, we sought to redirect an exocytic Rab, Sec4, onto endocytic vesicles by fusing the catalytic domain of the Sec4 GEF, Sec2, onto the CUE localization domain of Vps9, a GEF for the endocytic Rab Ypt51. The Sec2GEF-GFP-CUE construct localized to bright puncta predominantly near sites of polarized growth, and this localization was dependent on the ability of the CUE domain to bind to the ubiquitin moieties added to the cytoplasmic tails of proteins destined for endocytic internalization. Sec4 and Sec4 effectors were recruited to these puncta with various efficiencies. Cells expressing Sec2GEF-GFP-CUE grew surprisingly well and secreted protein at near-normal efficiency, implying that Golgi-derived secretory vesicles were delivered to polarized sites of cell growth despite the misdirection of Sec4 and its effectors. A low efficiency mechanism for localization of Sec2 to secretory vesicles that is independent of known cues might be responsible. In total, the results suggest that while Rabs may play a critical role in specifying the direction of vesicular transport, cells are remarkably tolerant of Rab misdirection.