Abstract

Significance PINK1 protein kinase and PARKIN UB ligase are mutated in inherited forms of Parkinson’s disease and several cancers. Thus, it is of great significance to understand normal functions that could be disrupted in disease. A role for PARKIN and PINK1 is in mediating autophagy of damaged mitochondria (mitophagy) through polyubiquitylation of numerous mitochondrial outer membrane proteins in a reaction that involves phosphorylation of both PARKIN and ubiquitin (UB) by PINK1. The mechanism remains unclear, however, due to challenges in defining individual steps in the pathway. Here, we use a UB replacement system to elucidate steps in the pathway that require PARKIN and/or UB phosphorylation by PINK1 and provide evidence of a PINK1- and UB-driven feed-forward mechanism important for efficient mitochondrial ubiquitylation and mitophagy.