A feed-forward pathway drives LRRK2 kinase membrane recruitment and apparent activation

Activating mutations in the Leucine Rich Repeat Kinase 2 (LRRK2) cause
Parkinson’s disease and activated LRRK2 phosphorylates a subset of Rab
GTPases. Moreover, Golgi-associated Rab29 can recruit LRRK2 to the surface
of the Golgi and activate it there for both auto- and Rab substrate
phosphorylation. Here we define the precise Rab29 binding region of the
LRRK2 Armadillo domain between residues 360-450 and show that this site,
termed “Site #1”, can also bind additional LRRK2 substrates, Rab8A and
Rab10. Moreover, we identify a distinct, N-terminal, higher affinity
interaction interface between LRRK2 phosphorylated Rab8 and Rab10 termed
“Site #2”, that can retain LRRK2 on membranes in cells to catalyze
multiple, subsequent phosphorylation events. Kinase inhibitor washout
experiments and mutation analysis demonstrate that rapid recovery of
kinase activity in cells depends on the ability of LRRK2 to associate with
phosphorylated Rab reaction products. Reconstitution of purified LRRK2
recruitment onto planar lipid bilayers decorated with Rab10 protein
demonstrates cooperative association of only active LRRK2 with
phospho-Rab10-containing membrane surfaces. These experiments reveal a
feed-forward pathway that provides spatial control and apparent membrane
activation of LRRK2 kinase activity.