Effect of p62/SQSTM1 polyubiquitination on its autophagic adaptor function and cellular survival under oxidative...

Oxidative stress induced by arsenite [As(III)] affects protein folding and results in increased levels of misfolded proteins or protein aggregates. Accumulation of misfolded protein aggregates may act as a cue signal for the oligomerization of the autophagic adaptor protein p62, which facilitates recognition of misfolded protein aggregates that are polyubiquitinated with K63 linkages. However, as the autophagic flux is impaired under exposure to As(III), p62 oligomers cannot be cleared by autophagy and accumulate as aggregates with Keap1. This results in the sequestration of Keap1 and the stabilization of Nrf2, which activates the non-canonical Nrf2-Keap1 pathway as an antioxidant response. In this study, we found that polyubiquitination of p62 itself increased upon exposure to As(III) to prevent further oligomerization of p62 and to increase the availability of functional free monomeric p62. We also found that monomeric p62 could also interact with ubiquitinated proteins and that the forced dimerization of p62 was sufficient to increase the interactions with ubiquitinated proteins, probably polyubiquitinated with K63 linkages. Upon exposure to As(III), (1) inability to form oligomeric p62 because of a mutation in the PB1 dimerization domain, or (2) reduced capability to generate monomeric p62 owing to diminished polyubiquitination of p62 itself, resulted in reduced viability of cells. Therefore, upon exposure to As(III), p62 initially needs to form oligomers to activate an antioxidant response pathway. Subsequently, p62 is polyubiquitinated to prevent further oligomerization and ensure the availability of free p62 monomers. We propose that the polyubiquitination of p62 under exposure to As(III) plays an important role in overcoming the impaired autophagic flux by regulating the oligomerization status of p62.