PDZD-8 interacts with LGG-2/LC3 to promote autolysosome formation

Abstract

LGG-2/LC3 and LGG-1/GABARAP are members of the Atg8 ubiquitin-like protein family, playing central roles in autophagy. Through a yeast two-hybrid screen, we identified the endoplasmic reticulum protein PDZD-8 as a novel interactor of the C. elegans autophagy protein LGG-2/LC3. We demonstrate that PDZD-8 binds specifically to LGG-2 and not LGG-1 via a canonical LC3-interacting region (LIR) and colocalizes with LGG-2-positive autophagosomes in vivo. Loss of PDZD-8 leads to a transient accumulation of enlarged LGG-2-positive autophagosomes that remain closely associated with lysosomes. Genetic disruption of core autophagy genes suppresses the pdzd-8 mutant phenotype, indicating that this phenotype is dependent on autophagy. Moreover, loss of rab-7, which blocks autophagosome-lysosome fusion, also suppresses the pdzd-8 phenotype, suggesting that the RAB-7-dependent fusion step is epistatic to PDZD-8 and that PDZD-8 acts downstream of autophagosome-lysosome maturation. Microscopy approaches (TEM, CLEM, and FIB-SEM) reveal an accumulation of fused autophagosome-lysosome structures, pointing to a transient block in autophagic flux in pdzd-8 mutants. Furthermore, PDZD-8 lacking either the SMP lipid-transfer domain or the LIR motif phenocopies the pdzd-8 null mutant, underscoring the importance of both domains. Our findings support a model in which PDZD-8, in conjunction with LGG-2/LC3, mediates ER-autophagosome/autolysosome contacts that are critical for subsequent autolysosome completion. Significance StatementAutophagy is a conserved cellular degradation pathway essential for maintaining homeostasis and responding to stress. Although the molecular machinery driving autophagosome formation and fusion with lysosomes is relatively well-characterized, the contribution of membrane contact sites to autophagic flux remains poorly understood. Here, we identify PDZD-8, an endoplasmic reticulum (ER) protein, as a novel interactor of the Caenorhabditis elegans LC3/Atg8 homolog LGG-2. We show that PDZD-8 regulates the completion of autolysosome maturation by bridging ER-autophagosome/autolysosome contacts. Loss of PDZD-8 results in the accumulation of enlarged autophagosomes adjacent to lysosomes and a transient block in autophagic flux, a phenotype that depends on the RAB-7-mediated tethering step. These findings reveal a previously unrecognized role for PDZD-8 in coordinating ER-autophagosome/autolysosome dynamics during autophagy.