Mutations in the leucine-rich repeat kinase 2 (LRRK2) are the most common known cause of PD. A major gap in our understanding of how LRRK2 causes PD is the lack of knowledge of LRRK2 physiological substrate(s). Thus, we propose to identify LRRK2 substrate(s) using a newly developed Protein Microarray technology with human proteins.
Increasing evidence suggests that LRRK2 is an upstream kinase in the signaling cascade that phosphorylates the downstream effectors (substrates). Thus, we will indentify LRRK2 substrate(s) by using a home-made high-throughput proteome chip with 4200 human proteins purified from a yeast expression system. We will then confirm the positive hits by various cell biology and biochemistry approaches.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
Identification of LRRK2 substrates will facilitate understanding of the molecular pathogenesis of LRRK2-related PD, provide potential innovative therapeutic targets, and help to develop the specific LRRK2 kinase assay and to screen for LRRK2 inhibitors to treat PD in future.
We anticipate that we will indentify LRRK2 substrate(s) from protein arrays and post confirmation experiments. If we identify a real substrate, it will lead to a series interesting projects for future studies: exploring the role of this substrate in PD pathogenesis and developing rational therape
Dr. Smith identified five possible candidate hits for LRRK2 substrates using a protein array. One protein in particular, a kinase, was shown to interact with G2019S-LRRK2 and to be primarily expressed within the cytoplasm. Dr. Smith is working to further validate this candidate protein.