Pink1 Kinase and Its Membrane Potential (Δψ)-dependent Cleavage Product Both Localize to Outer Mitochondrial Membrane by Unique Targeting Mode

Abstract

Background:The Parkinson-related kinase Pink1 is implicated in mitochondrial quality control. Results: Integration of multiple targeting signals results in the localization of Pink1 and its processing product to the surface of the outer membrane. Conclusion: Pink1 follows a unique import pathway that allows probing the functional integrity of mitochondria. Significance: Determining the Pink1 targeting mode provides the basis for defining its molecular function.

A Dimeric PINK1-containing Complex on Depolarized Mitochondria Stimulates Parkin Recruitment

“…First, we confirmed that exogenous and endogenous PINK1 form a complex following CCCP treatment (Fig. 1, A-C) (18,41). Thereafter, we identified components of this PINK1 complex.…”

“…5B). Complete degradation of PINK1 by proteinase K was similar to that of Tom20, suggesting that PINK1 WT and the pathogenic mutants localize to OMM (41,49).…”

“…Using immunoprecipitation in the presence of a crosslinker and with Tom20 and Tom22 under native conditions, Lazarou et al reported that PINK1 interacts with Tom20, Tom22, Tom40, and Tom70 (18). Meanwhile, Becker et al showed that an immunodetected Tom40 signal did not co-migrate with the PINK1 complex and thus concluded that the complex is unlikely to contain TOM components (41). Our data, which consist of two independent assays (the NAMOS assay and CN-PAGE), are consistent with the report by Lazarou et al (18), although we did not observe the incorporation of Tom70 (Fig.…”

“…Although two groups have reported that PINK1 forms a supermolecular complex after dissipation of ⌬⌿m (18,41), the incorporation of TOM components in the PINK1 complex was controversial. In general, the TOM complex consists of distinct import receptors (Tom20 or Tom70) and a core complex (common import receptor Tom22, pore Tom40, and organizer Tom5, -6, and -7).…”

“…When HeLa cells transiently expressing non-tagged PINK1 (ϳ60 kDa) were treated with CCCP to decrease ⌬⌿m, exogenous PINK1 formed a supermolecular complex (predicted size of ϳ850 kDa; referred to as 850-kDa complex hereafter) ( Fig. 1A) (18,41). We next used differential centrifugation to examine whether this complex is formed on mitochondria.…”