Lipidation-independent vacuolar functions of Atg8 rely on its noncanonical interaction with a vacuole membrane protein.

Xiao-Man Liu,Xiao-Min Du,Yoshinori Ohsumi,Akinori Yamasaki,Jian-Qiu Wu,Li-Lin Du,Nobuo N Noda,Valerie C Coffman,Hitoshi Nakatogawa

doi:10.7554/elife.41237

Abstract

The ubiquitin-like protein Atg8, in its lipidated form, plays central roles in autophagy. Yet, remarkably, Atg8 also carries out lipidation-independent functions in non-autophagic processes. How Atg8 performs its moonlighting roles is unclear. Here we report that in the fission yeast Schizosaccharomyces pombe and the budding yeast Saccharomyces cerevisiae, the lipidation-independent roles of Atg8 in maintaining normal morphology and functions of the vacuole require its interaction with a vacuole membrane protein Hfl1 (homolog of human TMEM184 proteins). Crystal structures revealed that the Atg8-Hfl1 interaction is not mediated by the typical Atg8-family-interacting motif (AIM) that forms an intermolecular β-sheet with Atg8. Instead, the Atg8-binding regions in Hfl1 proteins adopt a helical conformation, thus representing a new type of AIMs (termed helical AIMs here). These results deepen our understanding of both the functional versatility of Atg8 and the mechanistic diversity of Atg8 binding.

Highlights

Macroautophagy is an evolutionarily conserved bulk degradation pathway essential for cellular homeostasis
To identify Atg8-binding proteins in S. pombe, we performed affinity purification coupled with mass spectrometry analysis of C-terminally YFP-FLAG-His6 (YFH)-tagged Atg8 expressed in atg4D cells
We discovered the molecular underpinnings of the lipidation-independent vacuolar functions of Atg8 in yeasts

Summary

Introduction

Macroautophagy (hereafter autophagy) is an evolutionarily conserved bulk degradation pathway essential for cellular homeostasis. The structures showed that the Atg8-Hfl interaction is mediated by noncanonical mechanisms—using Hfl sequences adopting helical conformation (termed helical AIMs here) and in the case of budding yeast, involving a previously unreported binding site on Atg. The structures showed that the Atg8-Hfl interaction is mediated by noncanonical mechanisms—using Hfl sequences adopting helical conformation (termed helical AIMs here) and in the case of budding yeast, involving a previously unreported binding site on Atg8 These results unveil the molecular basis of the lipidationindependent vacuolar functions of Atg and expand our understanding of the diversity of Atg8-binding mechanisms

Results

Discussion

Materials and methods

Funding Funder