Methods for Assessing the Regulation of a Kinase by the Rab GTPase Ypt1

COPII coated vesicles that bud from the endoplasmic reticulum (ER) normally traffic to the Golgi. However, during starvation, COPII vesicles are redirected to the macroautophagy pathway where they become a membrane source for autophagosomes. Phosphorylation of the coat by the casein kinase 1 (CK1), Hrr25, is a prerequisite for vesicle uncoating and membrane fusion. CK1 family members were initially thought to be constitutively active kinases that are regulated through their subcellular localization. Recent studies, however, have shown that the Rab GTPase Ypt1 binds to and activates Hrr25 (CK1δ in mammals) to spatially regulate its kinase activity. Consistent with a direct role for Hrr25 in macroautophagy, hrr25and ypt1mutants are defective in autophagosome biogenesis. These studies have provided insights into how the itinerary of COPII vesicles is coordinated on two different trafficking pathways.