Deciphering the effect of LRRK2 constitutive phopshorylation and binding to 14-3-3 on protein function

Wednesday, October 10, 2012 — Poster Session II
Noon – 2:00 p.m	Natcher Conference Center, Building 45	NIA	CELLBIO-1

Authors

• R Chia
• A Beilina
• A Kaganovich
• M.R. Cookson

Abstract

Leucine rich repeat kinase 2 (LRRK2) is a large protein (~280kD) implicated in Parkinson’s disease (PD). LRRK2 has kinase and GTPase activity, is constitutively phosphorylated (CP) by another kinase and when phosphorylated, binds to 14-3-3 proteins. Inhibition of LRRK2 kinase activity resulted in loss of CP and 14-3-3 interaction. Here we investigated the effects of a LRRK2 inhibitor, IN1, on the biochemical and cellular properties of LRRK2. For kinase active, but not kinase dead (KD) mutants, IN1 treatment resulted in increased in GTP binding capacity, formation of large complexes >700kDa, and cytoplasmic relocalization of LRRK2 to filamentous structures. Interaction of LRRK2 with Hsp90 was independent of IN1, suggesting that loss of CP does not render LRRK2 unfolded but alters the interaction with other proteins, possibly with GTPase binding partners due to its increased GTP binding capacity. Interestingly, KD did not behave in a similar manner as kinase-inhibited LRRK2. Chronic expression of KD may result in the activation of a compensatory mechanism that may mask the phenotype observed in IN1 treated cells. In aggregate, these data suggest that changes in the physiological properties of LRRK2 occur in a kinase-dependent manner. This may be an important consideration when designing LRRK2-PD therapeutics.

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