Peroxisomes are important single membrane organelles which are required for certain metabolic pathways. In yeast, Pichia pastoris, peroxisomes are degraded by selective autophagy pathway termed pexophagy. Autophagy is process whereby proteins and organelles are delivered to the vacuole and degraded. Previous studies indicated Atg30 as the receptor for pexophagy, which interacts with the autophagic machinery (Atg11 and Atg17) as well as peroxisomal membrane (Pex3 and Pex14). However, unlike other selective autophagy receptors, Atg30 does not interact with Atg8, a major component of the isolation membrane that sequester peroxisomes during pexophagy. Here, we found the missing link, a novel peroxisomal membrane protein Rppa09976, which bridges peroxisomal (Pex3 and Atg30) and isolation (Atg8) membranes. Rppa09976 is phosphoprotein that contains an N-terminal acyl-CoA binding domain, three putative Atg8 binding sites and two transmembrane domains. Rppa09976 is expressed in methanol medium and phosphorylated upon arrival to peroxisomes from ER. We purified the Rppa09976-HA fusion protein and mapped 21 phosphorylation sites by mass spectrometry. The functional role of Rppa09976 phosphorylation awaits further studies. We showed that two of the three putative Atg8 binding sites at the N-terminal of the protein are required for Atg8 binding and pexophagy. Point mutation of the conserved acyl-CoA binding site also leads to a pexophagy defect. Hence, we propose that Rppa09976 provides acyl-CoA for the growing isolation membranes that sequester peroxisomes

Autophagy is a membrane trafficking pathway that sequesters proteins and organelles into autophagosomes. The selectivity of this pathway is determined by autophagy receptors, such as the Pichia pastoris autophagy-related protein 30 (Atg30), which controls the selective autophagy of peroxisomes (pexophagy) through the assembly of a receptor protein complex (RPC). However, how the pexophagic RPC is regulated for efficient formation of the phagophore, an isolation membrane that sequesters the peroxisome from the cytosol, is unknown. Here we describe a new, conserved acyl-CoA-binding protein, Atg37, that is an integral peroxisomal membrane protein required specifically for pexophagy at the stage of phagophore formation. Atg30 recruits Atg37 to the pexophagic RPC, where Atg37 regulates the recruitment of the scaffold protein, Atg11. Palmitoyl-CoA competes with Atg30 for Atg37 binding. The human orthologue of Atg37, acyl-CoA-binding domain containing protein 5 (ACBD5), is also peroxisomal and is required specifically for pexophagy. We suggest that Atg37/ACBD5 is a new component and positive regulator of the pexophagic RPC.