Oligomerization of p62 allows for selection of ubiquitinated cargo and isolation membrane during selective autophagy

Abstract

Autophagy is a major pathway for the clearance of harmful material from the cytoplasm. During autophagy, cytoplasmic material is delivered into the lysosomal system by organelles called autophagosomes. Autophagosomes form in a de novo manner and, in the course of their formation, isolate cargo material from the rest of the cytoplasm. Cargo specificity is conferred by autophagic cargo receptors that selectively link the cargo to the autophagosomal membrane decorated with ATG8 family proteins such as LC3B. Here we show that the human cargo receptor p62/SQSTM-1 employs oligomerization to stabilize its interaction with LC3B and linear ubiquitin when they are clustered on surfaces. Thus, oligomerization enables p62 to simultaneously select for the isolation membrane and the ubiquitinated cargo. We further show in a fully reconstituted system that the interaction of p62 with ubiquitin and LC3B is sufficient to bend the membrane around the cargo.DOI: http://dx.doi.org/10.7554/eLife.08941.001

Mechanisms of Selective Autophagy in Normal Physiology and Cancer

“…As binding of poly-ubiquitin to the UBA domain inhibits dephosphorylation, an increase in poly-ubiquitylated aggregates in the cell such as in the situation of proteasome overload or inhibition then shifts the balance of p62 to the phosphorylated form thereby enhancing aggrephagy[89]. Another intrinsic feature of p62 that contributes to aggrephagy is its ability to oligomerize via its PB1 domain thereby increasing the avidity of p62 for membrane surface clustered LC3B and ubiquitylated aggregates[79,90,91]. Oligomerization therefore links selection of phagophore and ubiquitylated cargo.…”

HSF1 stress response pathway regulates autophagy receptor SQSTM1/p62-associated proteostasis

“…report that SQSTM1 oligomerization is essential for its localization to the autophagosome formation site where SQSTM1 associates with upstream autophagy factors such as ULK1 and VMP1 41 . Moreover, the interaction of SQSTM1 with LC3 is promoted by SQSTM1 oligomerization 42 . Thus, the initiation of aggrephagy might be triggered by SQSTM1 oligomerization and localization to the autophagosome formation site after phosphorylation and substrate capturing of SQSTM1.…”

Mechanism of cargo-directed Atg8 conjugation during selective autophagy

“…Consistent with exclusion of Atg12~Atg5-Atg16 from the concave side of the isolation membrane, the Atg12~Atg5-Atg16 complex is excluded from the autophagosomal lumen in vivo (Mizushima et al, 2003). On the concave side Atg8 may be subsequently bound with high avidity by the cargo receptors, which could result in close apposition of the membrane and the cargo and thus exclusion of non-cargo material from the autophagosome (Figure 7) (Abert et al, 2016; Sawa-Makarska et al, 2014; Wurzer et al, 2015). As a consequence, this mechanism would localize the Atg8 conjugation machinery to the highly curved edge of the membrane where the isolation membrane has not yet formed, which may additionally stimulate Atg8 conjugation (Nath et al, 2014).…”

“…The formed complex was added to cells supplied with 2 ml fresh DMEM containing serum and antibiotics and incubated for two days. Thereafter co-transfection of plasmids containing a siRNA resistant p62 variant in pmCherry-C1 (SMC516, Wurzer et al, 2015) and Atg5 in pEGFP-C1 (SMC255) or pEGFP-C1 vector only was performed. 1 to 1.5 µg of plasmid-DNA were mixed with Fugene6 (Promega, WI, USA) in a 1 μg:3 μl ratio (DNA:Fugene6) in serum-free medium and incubated at RT for 15 to 45 min.…”