Transport of endo-β-mannosidase AtEBM from cytoplasm to the vacuole through autophagy pathway in Arabidopsis

Autophagy is a conserved eukaryotic intracellular degradation pathway that exhibits specialized forms, such as the biosynthetic cytoplasm-to-vacuole targeting (Cvt) pathway in yeast. This pathway selectively delivers several hydrolase precursors to the vacuole. While the core autophagy machinery is highly conserved between yeast and plants, the existence of a similar Cvt-like pathway for vacuolar hydrolase trafficking in plants remains unclear. Here, we found that Arabidopsis thaliana endo-{beta}-mannosidase (AtEBM), a glycosidase involved in degrading vacuolar N-glycans released from glycoproteins, is selectively transported to the vacuole via autophagy. Further analyses revealed that AtEBM directly interacts with the core autophagy protein ATG8 through an ATG8-interacting motif (AIM). This interaction is essential for sequestering AtEBM into autophagosomes. Remarkably, the AIM is conserved among EBM orthologs across the green plant lineage. In addition, several phenotypic studies showed that atebm mutants generated through CRISPR-Cas9 editing did not exhibit autophagy defects, including early senescence and hypersensitivity to nutrient starvation, but rather had reduced fecundity. Thus, these findings reveal a Cvt-like pathway in plants that fulfills a biosynthetic role by selectively delivering the glycosidase AtEBM to the vacuole.