ADP-ribosylation factor (ARF) and ARF-like (ARL) proteins are members of the ARF family, which are critical components of several different vesicular trafficking pathways. ARFs have little or no detectable GTPase activity without the assistance of a GTPase-activating protein (GAP). Here, we demonstrate that yeast Gcs1p exhibits GAP activity toward Arl1p and Arf1p in vitro, and Arl1p can interact with Gcs1p in a GTP-dependent manner. Arl1p was observed both on trans -Golgi and in cytosol and was recruited from cytosol to membranes in a GTP-dependent manner. In gcs1 mutant cells, the fraction of Arl1p in cytosol relative to trans -Golgi was less than it was in wild-type cells. Increasing Gcs1p levels returned the distribution toward that of wild-type cells. Both Arl1p and Gcs1p influenced the distribution of Imh1p, an Arl1p effector. Our data are consistent with the conclusion that Arl1p moves in a dynamic equilibrium between trans -Golgi and cytosol, and the release of Arl1p from membranes in cells requires the hydrolysis of bound GTP, which is accelerated by Gcs1p.
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