Loss of function mutations in PINK1, a mitochondrial kinase, are associated with recessive parkinsonism. Previous work has shown that the kinase activity of PINK1 is likely to be critically important in the neuroprotective function of PINK1. Furthermore, work originating from Drosophila models but subsequently confirmed in cell culture systems suggests that PINK1 works via the E3 ligase parkin, but the mechanistic basis of this observation is unclear. We have made and previously published a series of human neuroblastoma cell lines from the M17 background where we used lentiviruses to stably integrate vectors to either increase or decrease (using shRNA) PINK1 expression. These lines were used by Richard Youles laboratory in the National Institute of Neurological Diseases and Stroke to show that active PINK1 kinase is required for the recruitment of parkin to the mitochondrial membrane under conditions of mitochondrial damage. The phenotype of parkin relocalization is therefore a marker for the activation of the PINK1 kinase at the mitochondrial membrane. We are currently following up this observation by examining whether other agents that damage mitochondria will have the same effect and using high throughput strategies attempting to identify other components of this pathway.
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