Abstract
Autophagic receptor p62 is a critical mediator of cell detoxification, stress response, and metabolic programs and is commonly deregulated in human diseases. The diverse functions of p62 arise from its ability to interact with a large set of ligands, such as arginylated (Nt-R) substrates. Here, we describe the structural mechanism for selective recognition of Nt-R by the ZZ domain of p62 (p62ZZ). We show that binding of p62ZZ to Nt-R substrates stimulates p62 aggregation and macroautophagy and is required for autophagic targeting of p62. p62 is essential for mTORC1 activation in response to arginine, but it is not a direct sensor of free arginine in the mTORC1 pathway. We identified a regulatory linker (RL) region in p62 that binds p62ZZ in vitro and may modulate p62 function. Our findings shed new light on the mechanistic and functional significance of the major cytosolic adaptor protein p62 in two fundamental signaling pathways.
Highlights
Autophagic receptor p62 is a critical mediator of cell detoxification, stress response, and metabolic programs and is commonly deregulated in human diseases
We report the structural basis underlying specific recognition of Nt-R substrates by the ZZ domain of p62. We demonstrate that this interaction promotes p62 aggregation and is necessary for macroautophagy
To quantitatively monitor functional autophagy, autolysosome formation in cells, an mCherry-GFP-LC3 tandem construct was stably expressed, and the acidic quenching of green fluorescent protein (GFP) signal relative to the stable expression of mCherry was measured by flow cytometry[22] (Fig. 4a)
Summary
Autophagic receptor p62 is a critical mediator of cell detoxification, stress response, and metabolic programs and is commonly deregulated in human diseases. Dissociation constant (Kd) measured by microscale thermophoresis (MST) for the interaction of p62ZZ with REEE was found to be 5.6 μM, whereas binding of the RAEE peptide was reduced (Kd = 14 μM) Binding of p62ZZ to arginylated substrates induces autophagy.
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