In eukaryotic cells, the endomembrane system consists of multiple membrane-bound organelles, which play essential roles in the precise transportation of various cargo proteins. In plant cells, vacuoles are regarded as the terminus of catabolic pathways whereas the selection and transport of vacuolar cargoes are mainly mediated by two types of organelles, multivesicular bodies (MVBs) also termed prevacuolar compartments (PVCs) and autophagosomes. MVBs are single-membrane bound organelles with intraluminal vesicles and mediate the transport between the trans-Golgi network (TGN) and vacuoles, while autophagosomes are double-membrane bound organelles, which mediate cargo delivery to the vacuole for degradation and recycling during autophagy. Great progress has been achieved recently in identification and characterization of the conserved and plant-unique regulators involved in the MVB and autophagosome pathways. In this review, we present an update on the current knowledge of these key regulators and pay special attention to their conserved protein domains. In addition, we discuss the possible interplay between the MVB and autophagosome pathways in regulating vacuolar degradation in plants.