The entry of unclosed autophagosomes into vacuoles and its physiological relevance

Abstract

It is widely stated in the literature that closed mature autophagosomes (APs) fuse with lysosomes/vacuoles during macroautophagy/autophagy. Previously, we showed that unclosed APs accumulated as clusters outside vacuoles in Vps21/Rab5 and ESCRT mutants after a short period of nitrogen starvation. However, the fate of such unclosed APs remains unclear. In this study, we used a combination of cellular and biochemical approaches to show that unclosed double-membrane APs entered vacuoles and formed unclosed single-membrane autophagic bodies after prolonged nitrogen starvation or rapamycin treatment. Vacuolar hydrolases, vacuolar transport chaperon (VTC) proteins, Ypt7, and Vam3 were all involved in the entry of unclosed double-membrane APs into vacuoles in Vps21-mutant cells. Overexpression of the vacuolar hydrolases, Pep4 or Prb1, or depletion of most VTC proteins promoted the entry of unclosed APs into vacuoles in Vps21-mutant cells, whereas depletion of Pep4 and/or Prb1 delayed the entry into vacuoles. In contrast to the complete infertility of diploid cells of typical autophagy mutants, diploid cells of Vps21 mutant progressed through meiosis to sporulation, benefiting from the entry of unclosed APs into vacuoles after prolonged nitrogen starvation. Overall, these data represent a new observation that unclosed double-membrane APs can enter vacuoles after prolonged autophagy induction, most likely as a survival strategy.

Characterization of Aspartic Proteases from Paracoccidioides brasiliensis and Their Role in Fungal Thermo-Dimorphism

“…Finally, an evaluation of the interactome of PbSap1, PbSap2, PbYap1, and PbYap2 was performed and suggested that the function of these proteases is involved in the regulation and acquisition of carbon and nitrogen, autophagy, nucleic acid metabolisms, and the activation of other proteases. Particularly, the interaction of aspartic proteases Pep4 and Atg8 plays an important role in autophagy, since the absence of Pep4 (Vacuolar protease A) completely blocks autophagy [ 73 ].…”

Phagophore closure, autophagosome maturation and autophagosome fusion during macroautophagy in the yeast Saccharomyces cerevisiae

“…It is very difficult to reconcile these discrepancies, but it may suggest that the ESCRT complexes are not absolutely required for phagophore closure under specific conditions and/or in certain strain backgrounds. Conversely, it cannot be excluded that unclosed autophagosomes fuse with the vacuole in certain situations [33]. Moreover, some of the phenotypes observed in ESCRT mutants could be due to indirect defects caused by an impairment of the endosomal trafficking of key autophagy players such as Atg9 that get missorted in class E compartments, which are aberrant late endosomal structures present in these cells [34].…”

Autophagy beyond convention : plant-specific mechanisms for cellular recycling

“…Sealing of phagophores was previously considered extremely crucial since unsealed autophagosomes cannot fuse with the lytic compartment. However, a recent study in yeast demonstrates that unsealed autophagosomes can enter the vacuoles upon prolonged autophagy induction (Wu et al, 2022). It remains to be determined whether this adaptation is exclusive to yeast or if it can also occur in plants and mammals.…”