Interaction between parkin and alpha‐synuclein

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Mutations in the parkin gene have been identified in autosomal recessive Parkinson's disease (ARPD) throughout the world. Several groups have shown that parkin acts as an ubiquitin (Ub) ligase in in vivo and in vitro studies and identified several molecules to be ubiquitinated by parkin. Two rare mutations, Ala53Thr and Ala30Pro, in the α‐synuclein (αS) gene have been identified in autosomal dominant PD cases. αS is a major constituent of Lewy bodies the pathological hallmark of PD. Therefore, αvS metabolism is likely to play a central role in the pathogenesis of PD. We investigated the possible interaction between the two familial PD‐linked gene products, parkin and αS. We first demonstrated that parkin epitopes can be found within Lewy bodies by immunohistochemistry and immuno‐EM. We also detected parkin biochemically in affinity purified Lewy bodies. Furthermore, we identified a protein complex in normal human brain that includes parkin as an E3 Ub ligase, UbcH7 as its associated E2 protein and a novel 22 kDa glycosylated isoform of αS (αSp22) as its substrate. In an in vitro ubiquitination assay, αSp22 was further modified by normal but not mutant parkin into high Mr polyubiquitinated species. We observed accumulation of nonubiquitinated αSp22 in ARPD brain homogenates. Importantly enzymatic digestion of αSp22 abolish the binding of recombinant parkin with deglycosylated αS. We conclude that αSp22 is a substrate for parkin's Ub ligase activity in normal brain, and that loss of function in parkin‐deficient ARPD causes pathological αSp22 accumulation. These findings demonstrate a critical functional relationship between the two PD‐linked gene products.