The entry of unclosed autophagosomes into vacuoles and its physiological relevance

PLoS Genet. 2022 Oct 13;18(10):e1010431. doi: 10.1371/journal.pgen.1010431. eCollection 2022 Oct.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Autophagosomes / metabolism
  • Autophagy / genetics
  • Endosomal Sorting Complexes Required for Transport / metabolism
  • Hydrolases / metabolism
  • Molecular Chaperones / metabolism
  • Nitrogen / metabolism
  • Saccharomyces cerevisiae / metabolism
  • Saccharomyces cerevisiae Proteins* / metabolism
  • Sirolimus / metabolism
  • Sirolimus / pharmacology
  • Vacuoles* / genetics
  • Vacuoles* / metabolism
  • rab GTP-Binding Proteins / metabolism

Substances

  • Endosomal Sorting Complexes Required for Transport
  • Molecular Chaperones
  • Saccharomyces cerevisiae Proteins
  • Hydrolases
  • VPS21 protein, S cerevisiae
  • YPT7 protein, S cerevisiae
  • rab GTP-Binding Proteins
  • Nitrogen
  • Sirolimus

Grants and funding

This work was supported by grants from the Natural Science Foundation of China grant numbers 91954125, 31871428, 31671479 to Y. L. (https://isisn.nsfc.gov.cn), the National Key R&D Program of China grant number 2017YFA0504700 to Y. X. (https://service.most.gov.cn/), and the open funding from the State Key Laboratory of Microbial Metabolism, Shanghai Jiao Tong University grant number MMLKF21-06 to Y. L. & Z. X. (https://skmml.sjtu.edu.cn/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.