P62/SQSTM1 is a novel leucine-rich repeat kinase 2 (LRRK2) substrate that enhances neuronal toxicity

Area of Science:

• Neuroscience
• Genetics
• Molecular Biology

Background:

• Autosomal-dominant mutations in leucine-rich repeat protein kinase 2 (LRRK2) are a common genetic cause of Parkinson's disease (PD).
• Increased LRRK2 kinase activity in pathogenic mutations suggests substrate hyperphosphorylation contributes to PD pathogenesis.
• Identifying LRRK2 downstream targets is critical for understanding disease mechanisms.

Purpose of the Study:

• To identify novel endogenous interacting partners and substrates of LRRK2.
• To investigate the role of p62/SQSTM1 as a substrate of LRRK2.
• To explore the impact of LRRK2 mutations and inhibitors on p62 phosphorylation and neurotoxicity.

Main Methods:

• Mass spectrometry and phospho-specific antibodies were used to identify and quantify LRRK2-mediated p62 phosphorylation.
• Experiments were conducted both in vitro and in cell models expressing various LRRK2 mutations.
• The effect of LRRK2 inhibitors and specific protein domains on phosphorylation was assessed.

Main Results:

• The signaling adapter protein p62/SQSTM1 was identified as a novel LRRK2 interacting partner and substrate, phosphorylated at Thr138.
• Pathogenic LRRK2 mutations (N1437H, R1441C/G/H, Y1699C, G2019S) increased p62 phosphorylation, while I2020T and G2385R showed decreased phosphorylation.
• LRRK2 inhibitor treatment blocked p62 phosphorylation, and LRRK2's amino-terminus and p62's ubiquitin-binding domain were crucial for this interaction.

Conclusions:

• p62 is a novel substrate of LRRK2, and its phosphorylation by LRRK2 regulates neurotoxic pathways.
• LRRK2-mediated p62 phosphorylation represents a novel signaling node in Parkinson's disease.
• p62 phosphorylation can serve as a pharmacodynamic marker for LRRK2 kinase activity and inhibitor efficacy.