Enhancement of ubiquitination-dependent mitophagy by unconventionally-acting PROTACs

Enhancing mitophagy, a naturally-occurring cellular process for elimination of damaged mitochondria, holds great promise for the intervention of many human diseases. Proteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules that induce ubiquitination and subsequent proteasome-mediated degradation of a target protein through simultaneously binding to the target protein and an E3 ubiquitin ligase. However, the narrow cavity of the proteasome prevents the degradation of mitochondria. Here we show that the E3 ubiquitin ligase MAP3K1, when recruited to the outer mitochondria membrane (OMM) protein TSPO or VDAC by our PROTAC-designed molecules (termed “mitophagy-enhancing chimeras”, or MECs), induced extensive K63 ubiquitination of TSPO and other OMM proteins, reminiscent of the PINK1-activated PARKIN, without triggering proteasome-mediated degradation of TSPO. Aided by NBR1 and Nur77, this increased K63 ubiquitination of OMM proteins triggered mitophagy exclusively for damaged mitochondria, leading to improved mitochondria function and diminished cellular ROS. With the capability to enhance mitophagy at low nanomolar concentrations, MECs effectively inhibited NLRP3 inflammasome activation, abrogated acetaminophen-induced acute liver injury and mitigated high-fat diet-induced obesity in mice. Our work provided a proof-of-concept for developing unconventionally-acting PROTACs to achieve degradation of damaged mitochondria and possibly other organelles.