Three overlapping pathways mediate the transport of cytoplasmic material to the 
vacuole in Saccharomyces cerevisiae. The cytoplasm to vacuole targeting (Cvt) 
pathway transports the vacuolar hydrolase, aminopeptidase I (API), whereas 
pexophagy mediates the delivery of excess peroxisomes for degradation. Both the 
Cvt and pexophagy pathways are selective processes that specifically recognize 
their cargo. In contrast, macroautophagy nonselectively transports bulk cytosol 
to the vacuole for recycling. Most of the import machinery characterized thus 
far is required for all three modes of transport. However, unique features of 
each pathway dictate the requirement for additional components that 
differentiate these pathways from one another, including at the step of specific 
cargo selection.We have identified Cvt9 and its Pichia pastoris counterpart 
Gsa9. In S. cerevisiae, Cvt9 is required for the selective delivery of precursor 
API (prAPI) to the vacuole by the Cvt pathway and the targeted degradation of 
peroxisomes by pexophagy. In P. pastoris, Gsa9 is required for glucose-induced 
pexophagy. Significantly, neither Cvt9 nor Gsa9 is required for 
starvation-induced nonselective transport of bulk cytoplasmic cargo by 
macroautophagy. The deletion of CVT9 destabilizes the binding of prAPI to the 
membrane and analysis of a cvt9 temperature-sensitive mutant supports a direct 
role of Cvt9 in transport vesicle formation. Cvt9 oligomers peripherally 
associate with a novel, perivacuolar membrane compartment and interact with 
Apg1, a Ser/Thr kinase essential for both the Cvt pathway and autophagy. In P. 
pastoris Gsa9 is recruited to concentrated regions on the vacuole membrane that 
contact peroxisomes in the process of being engulfed by pexophagy. These 
biochemical and morphological results demonstrate that Cvt9 and the P. pastoris 
homologue Gsa9 may function at the step of selective cargo sequestration.