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Analysis of ATG4C function in vivo.

Isaac Tamargo-Gómez1,2,3, Gemma G Martínez-García1,2,3, María F Suárez3

  • 1Departamento de Biología Funcional, Facultad de Medicina, Universidad de Oviedo, Oviedo, Spain.

Autophagy
|July 17, 2023
PubMed
Summary

Autophagy-related 4 (ATG4) proteases cleave ATG8 family proteins, essential for autophagy. This study investigates the specific roles of ATG4A, ATG4B, ATG4C, and ATG4D in cleaving ATG8 proteins like MAP1LC3B/LC3B and GABARAPL1.

Keywords:
Animal modelsGABARAPLC3autophagyfibrosarcomalymphocyte

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Autophagy is a crucial cellular process for degrading and recycling damaged components.
  • ATG8 family proteins (e.g., MAP1LC3/LC3, GABARAP) are key players in autophagosome formation.
  • ATG4 proteases are essential for processing ATG8 proteins, but their specific substrate preferences and functions remain incompletely understood.

Purpose of the Study:

  • To elucidate the distinct roles of the four mammalian ATG4 proteases (ATG4A, ATG4B, ATG4C, ATG4D).
  • To determine the specific ATG8 family proteins (including MAP1LC3A/LC3A, MAP1LC3B/LC3B, GABARAP, GABARAPL1, GABARAPL2) cleaved by each ATG4 paralog.
  • To investigate the impact of ATG4 activity on lipidation of ATG8 proteins with phosphatidylethanolamine (PE).

Main Methods:

  • In vitro protease assays using recombinant ATG4 proteins and ATG8 substrates.
  • Cell-based assays to assess ATG8 processing and lipidation in cells with depleted or overexpressed ATG4s.
  • Mass spectrometry to identify cleavage sites and modified ATG8 proteins.

Main Results:

  • Differential substrate specificity was observed among ATG4 paralogs for ATG8 proteins.
  • ATG4B and ATG4D showed significant activity towards MAP1LC3B/LC3B and GABARAPL1.
  • ATG4A and ATG4C exhibited distinct cleavage patterns and influenced PE conjugation of specific ATG8 proteins.

Conclusions:

  • Mammalian ATG4 proteases possess specialized functions in cleaving ATG8 proteins, contributing to the fine-tuning of autophagy.
  • The specific ATG4-ATG8 interactions regulate key steps in autophagosome biogenesis and cargo recognition.
  • Understanding these specificities is critical for deciphering autophagy regulation in health and disease.