Plant NBR1 is a selective autophagy substrate and a functional hybrid of the mammalian autophagic adapters NBR1 and p62/SQSTM1

Autophagy. 2011 Sep;7(9):993-1010. doi: 10.4161/auto.7.9.16389. Epub 2011 Sep 1.

Abstract

(Macro)autophagy encompasses both an unselective, bulk degradation of cytoplasmic contents as well as selective autophagy of damaged organelles, intracellular microbes, protein aggregates, cellular structures and specific soluble proteins. Selective autophagy is mediated by autophagic adapters, like p62/SQSTM1 and NBR1. p62 and NBR1 are themselves selective autophagy substrates, but they also act as cargo receptors for degradation of other substrates. Surprisingly, we found that homologs of NBR1 are distributed throughout the eukaryotic kingdom, while p62 is confined to the metazoans. As a representative of all organisms having only an NBR1 homolog we studied Arabidopsis thaliana NBR1 (AtNBR1) in more detail. AtNBR1 is more similar to mammalian NBR1 than to p62 in domain architecture and amino acid sequence. However, similar to p62, AtNBR1 homo-polymerizes via the PB1 domain. Hence, AtNBR1 has hybrid properties of mammalian NBR1 and p62. AtNBR1 has 2 UBA domains, but only the C-terminal UBA domain bound ubiquitin. AtNBR1 bound AtATG8 through a conserved LIR (LC3-interacting region) motif and required co-expression of AtATG8 or human GABARAPL2 to be recognized as an autophagic substrate in HeLa cells. To monitor the autophagic sequestration of AtNBR1 in Arabidopsis we made transgenic plants expressing AtNBR1 fused to a pH-sensitive fluorescent tag, a tandem fusion of the red, acid-insensitive mCherry and the acid-sensitive yellow fluorescent proteins. This strategy allowed us to show that AtNBR1 is an autophagy substrate degraded in the vacuole dependent on the polymerization property of the PB1 domain and of expression of AtATG7. A functional LIR was required for vacuolar import.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / chemistry
  • Adaptor Proteins, Signal Transducing / metabolism*
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Arabidopsis / metabolism*
  • Arabidopsis Proteins / chemistry
  • Arabidopsis Proteins / metabolism*
  • Autophagy*
  • Autophagy-Related Protein 8 Family
  • Carrier Proteins / chemistry
  • Carrier Proteins / metabolism*
  • Conserved Sequence
  • Cytosol / metabolism
  • Eukaryotic Cells / metabolism
  • Evolution, Molecular
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins
  • Microfilament Proteins / metabolism
  • Models, Molecular
  • Molecular Sequence Data
  • Phylogeny
  • Plant Cells / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Protein Transport
  • Proteins / chemistry
  • Proteins / metabolism*
  • Sequence Homology, Amino Acid
  • Sequestosome-1 Protein
  • Ubiquitin / metabolism
  • Vacuoles / metabolism

Substances

  • Adaptor Proteins, Signal Transducing
  • Arabidopsis Proteins
  • Autophagy-Related Protein 8 Family
  • Carrier Proteins
  • GABARAPL2 protein, human
  • Intracellular Signaling Peptides and Proteins
  • Microfilament Proteins
  • NBR1 protein, Arabidopsis
  • NBR1 protein, human
  • Proteins
  • SQSTM1 protein, human
  • Sequestosome-1 Protein
  • Ubiquitin