A molecular explanation for the recessive nature of parkin-linked Parkinson’s disease

Abstract

Mutations in the park2 gene, encoding the RING-inBetweenRING-RING E3 ubiquitin ligase parkin, cause 50% of autosomal recessive juvenile Parkinsonism cases. More than 70 known pathogenic mutations occur throughout parkin, many of which cluster in the inhibitory amino-terminal ubiquitin-like domain, and the carboxy-terminal RING2 domain that is indispensable for ubiquitin transfer. A structural rationale showing how autosomal recessive juvenile Parkinsonism mutations alter parkin function is still lacking. Here we show that the structure of parkin RING2 is distinct from canonical RING E3 ligases and lacks key elements required for E2-conjugating enzyme recruitment. Several pathogenic mutations in RING2 alter the environment of a single surface-exposed catalytic cysteine to inhibit ubiquitination. Native parkin adopts a globular inhibited conformation in solution facilitated by the association of the ubiquitin-like domain with the RING-inBetweenRING-RING C-terminus. Autosomal recessive juvenile Parkinsonism mutations disrupt this conformation. Finally, parkin autoubiquitinates only in cis, providing a molecular explanation for the recessive nature of autosomal recessive juvenile Parkinsonism.

Synergistic recruitment of UbcH7~Ub and phosphorylated Ubl domain triggers parkin activation

“…We noted that some of the largest chemical shift changes were in this region of the tether (Y391, R392, D394), which undergoes multiple rearrangements in crystal structures of parkin (Kumar et al , 2017). NMR dynamics experiments (Fig EV1; Kumar et al , 2015) and a structure of the isolated IBR–tether–RING2(Rcat) region (Spratt et al , 2013) indicate this region is very flexible and likely adopts multiple conformations in solution. We interpreted the chemical shift changes within the tether region to result from both direct E2 binding and an altering of the tether position to accommodate the E2 enzyme.…”

“…We first analysed the observed ubiquitin surface involved at the Parkin:UbcH7‐Ub interface using distinct assays that monitor transfer of ubiquitin from the E2 onto parkin and subsequently onto the substrate. To assess UbcH7 mediated ubiquitin loading of parkin, we generated a pParkin RING2(Rcat) mutant (C431S + H433A, referred to as pParkin CH ) that is able to trap an E3~Ub oxyester intermediate (Spratt et al , 2013; Kumar et al , 2017). We rationalized that ubiquitin loading to pParkin CH would depend on the interaction of Ub in the UbcH7~Ub conjugate with pParkin.…”

“…Full‐length human parkin (1–465), Ubl (1–76), R0RBR (141–465), Drosophila melanogaster RING2 (410–482) and other parkin variants were expressed and purified as described previously (Chaugule et al , 2011; Spratt et al , 2013). Briefly, His‐smt3‐parkin constructs were expressed in BL21(DE3) cells at 37°C to an OD 600 of 0.8.…”

Control of developmental networks by Rac/Rho small GTPases: How cytoskeletal changes during embryogenesis are orchestrated

“…PDR‐1/Parkin maintains CED‐10/Rac levels to prevent over‐activated apoptotic cell engulfment or abnormal distal tip cell (DTC) migration 72. In humans, autosomal recessive juvenile Parkinson's disease (AR‐JP) is characterized by parkin mutations 76, a possible mechanism underlying this or other neurodegenerative disease characterized by abnormal CED‐10/Rac degradation could be an acceleration of neurons phagocytosis. Further studies are still needed to test the relevance of Rac‐1‐Parkin interaction in diseases.…”

Phosphorylated ubiquitin: a new shade of PINK1 in Parkin activation

“…The kinase activity of PINK1 is required for the translocation and activation of Parkin, a cytosolic E3 ubiquitin ligase that acts as a quality control guard by ubiquitinating proteins of the outer mitochondrial membrane to trigger selective autophagy of the damaged mitochondria, a process termed mitophagy ( Figure 1). Recent structures of Parkin revealed that the protein adopts an inactive conformation under basal conditions and needs to undergo a structural rearrangement to become active [2][3][4][5]. Mutation of residues involved in maintaining the inactive conformation (e.g., a tryptophan in the repressor element of Parkin, or REP) led to an increase in Parkin activity [2].…”