p62/SQSTM1 binds directly to Atg8/LC3 to facilitate degradation of ubiquitinated protein aggregates by autophagy.
Protein degradation by basal constitutive autophagy is important to avoid accumulation of polyubiquitinated protein aggregates and development of neurodegenerative diseases. The polyubiquitin-binding protein p62/SQSTM1 is degraded by autophagy. It is found in cellular inclusion bodies together with polyubiquitinated proteins and in cytosolic protein aggregates that accumulate in various chronic, toxic, and degenerative diseases. Here we show for the first time a direct interaction between p62 and the autophagic effector proteins LC3A and -B and the related gamma-aminobutyrate receptor-associated protein and gamma-aminobutyrate receptor-associated-like proteins. The binding is mediated by a 22-residue sequence of p62 containing an evolutionarily conserved motif. To monitor the autophagic sequestration of p62- and LC3-positive bodies, we developed a novel pH-sensitive fluorescent tag consisting of a tandem fusion of the red, acid-insensitive mCherry and the acid-sensitive green fluorescent proteins. This approach revealed that p62- and LC3-positive bodies are degraded in autolysosomes. Strikingly, even rather large p62-positive inclusion bodies (2 microm diameter) become degraded by autophagy. The specific interaction between p62 and LC3, requiring the motif we have mapped, is instrumental in mediating autophagic degradation of the p62-positive bodies. We also demonstrate that the previously reported aggresome-like induced structures containing ubiquitinated proteins in cytosolic bodies are dependent on p62 for their formation. In fact, p62 bodies and these structures are indistinguishable. Taken together, our results clearly suggest that p62 is required both for the formation and the degradation of polyubiquitin-containing bodies by autophagy.
Pankiv
Serhiy
S
Biochemistry Department, Institute of Medical Biology, University of Tromsø, 9037 Tromsø, Norway.
Clausen
Terje Høyvarde
TH
Lamark
Trond
T
Brech
Andreas
A
Bruun
Jack-Ansgar
JA
Outzen
Heidi
H
Øvervatn
Aud
A
Bjørkøy
Geir
G
Johansen
Terje
T
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000595:Amino Acid Sequence; D001343:Autophagy; D003600:Cytosol; D005456:Fluorescent Dyes; D006367:HeLa Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D046912:Multiprotein Complexes; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000494:pharmacology; D000071456:Sequestosome-1 Protein; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D005456:Fluorescent Dyes; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D046912:Multiprotein Complexes; D034741:RNA, Small Interfering; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D025801:Ubiquitin