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Related Experiment Video

Updated: Jul 18, 2025

Study of Protein-protein Interactions in Autophagy Research

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Published on: September 9, 2017

TOR-mediated Ypt1 phosphorylation regulates autophagy initiation complex assembly.

Weijing Yao¹, Yuting Chen¹, Yingcong Chen¹

¹Department of Biochemistry, and Department of Hepatobiliary and Pancreatic Surgery of the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

The EMBO Journal

|August 28, 2023

Summary

The Rab GTPase Ypt1 regulates autophagy initiation by controlling the stepwise assembly of ATG proteins. Its phosphorylation by TOR and interactions with Atg23 and Atg17 are crucial for autophagosome biogenesis.

Keywords:

ATG proteins TOR Ypt1 autophagy stepwise assembly

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Area of Science:

Cell Biology
Molecular Biology

Background:

Autophagy is a vital cellular process for degrading damaged components.
The precise molecular mechanisms governing the initiation of autophagy, particularly the assembly of autophagy-related (ATG) proteins, are not fully understood.
Rab GTPase Ypt1 (also known as Rab1) is a known regulator of autophagy.

Purpose of the Study:

To elucidate the role of Ypt1 in the stepwise assembly of ATG proteins during autophagy initiation.
To identify Ypt1 binding partners involved in autophagy complex formation.
To investigate the impact of Ypt1 phosphorylation by TOR on autophagy regulation.

Main Methods:

Co-immunoprecipitation assays to identify Ypt1 binding partners.
Analysis of ATG protein complex assembly and localization using genetic mutations and microscopy.
Site-directed mutagenesis to study the effect of Ypt1 phosphorylation on autophagy.

Main Results:

Atg23 and Atg17 were identified as direct binding partners of Ypt1, essential for stepwise assembly of autophagy initiation complexes.
Disruption of Ypt1-Atg23 interaction impaired Atg9 recruitment to the phagophore assembly site (PAS).
Ypt1 phosphorylation by TOR at Ser174 is a critical regulatory event; non-phosphorylatable Ypt1 enhances autophagy, while a phosphomimic mutant inhibits it.

Conclusions:

Ypt1 acts as a multifunctional assembly factor in autophagy initiation.
Ypt1's interactions with Atg23 and Atg17, and its TOR-mediated phosphorylation, are key to regulating ATG protein assembly and autophagosome biogenesis.
TOR-mediated Ypt1 phosphorylation provides a critical control point for autophagy initiation.