Rab GTPases are key regulators of membrane traffi c. The Rab GTPase Ypt1 is essential for endoplasmic reticulum (ER)-Golgi traffi c, intra-Golgi traffi c, and the macroautophagy pathway. To identify effectors on the macroautophagy pathway, known autophagy-related genes (Atg genes) required for macroautophagy were tagged with GFP and screened for mislocalization in the ypt1 - 2 mutant. At the pre-autophagosomal structure (PAS), the localization of the serine/threonine kinase Atg1 was affected in the ypt1 - 2 mutant. We then used an in vitro binding assay to determine if Atg1 and Ypt1 physically interact with each other and co-immunoprecipitation experiments were performed to address if Atg1 preferentially interacts with the GTP-bound form of Ypt1.
CITATION STYLE
Wang, J., Cervantes, S., Davis, S., & Ferro-Novick, S. (2015). Identifying a rab effector on the macroautophagy pathway. Methods in Molecular Biology, 1298, 117–125. https://doi.org/10.1007/978-1-4939-2569-8_10
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