1. Academic Validation
  2. PFKP facilitates ATG4B phosphorylation during amino acid deprivation-induced autophagy

PFKP facilitates ATG4B phosphorylation during amino acid deprivation-induced autophagy

  • Cell Signal. 2021 Jun;82:109956. doi: 10.1016/j.cellsig.2021.109956.
Xiuzhi Li 1 Lingling Sun 1 Guokai Yan 1 Xianghua Yan 2
Affiliations

Affiliations

  • 1 State Key Laboratory of Agricultural Microbiology, College of Animal Sciences and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, Hubei 430070, China; Hubei Provincial Engineering Laboratory for Pig Precision Feeding and Feed Safety Technology, Wuhan, Hubei 430070, China.
  • 2 State Key Laboratory of Agricultural Microbiology, College of Animal Sciences and Technology, Huazhong Agricultural University, Wuhan, Hubei 430070, China; The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, Hubei 430070, China; Hubei Provincial Engineering Laboratory for Pig Precision Feeding and Feed Safety Technology, Wuhan, Hubei 430070, China. Electronic address: xhyan@mail.hzau.edu.cn.
Abstract

ATG4B facilitates Autophagy by promoting autophagosome maturation through the reversible lipidation and delipidation of LC3. Recent reports have shown that phosphorylation of ATG4B regulates its activity and LC3 processing, leading to modulate Autophagy activity. However, the mechanism about how ATG4B phosphorylation is involved in amino acid deprivation-induced Autophagy is unclear. Here, we combined the tandem affinity purification with mass spectrometry (MS) and identified the ATG4B-interacting proteins including its well-known partner gamma-aminobutyric acid receptor-associated protein (GABARAP, a homolog of LC3) and phosphofructokinase 1 platelet isoform (PFKP). Further immunoprecipitation assays showed that amino acid deprivation strengthened the interaction between ATG4B and PFKP. By genetic depletion of PFKP using CRISPR/Cas9, we uncovered that PFKP loss reduced the degradation of LC3-II and p62 due to a partial block in autophagic flux. Furthermore, MS analysis of Flag-tagged ATG4B immunoprecipitates identified phosphorylation of ATG4B serine 34 residue (S34) and PFKP serine 386 residue (S386) under amino acid deprivation condition. In vitro kinase assay validated that PFKP functioning as a protein kinase phosphorylated ATG4B at S34. This phosphorylation could enhance ATG4B activity and p62 degradation. In addition, PFKP S386 phosphorylation was important to ATG4B S34 phosphorylation and Autophagy in HEK293T cells. In brief, our findings describe that PFKP, a rate-limiting Enzyme in the glycolytic pathway, functions as a protein kinase for ATG4B to regulate ATG4B activity and Autophagy under amino acid deprivation condition.

Keywords

ATG4B; Amino acid deprivation; Autophagy; PFKP; Phosphorylation.

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