Identification of protein phosphatase 1 as a regulator of the LRRK2 phosphorylation cycle.
A cluster of phosphorylation sites in LRRK2 (leucine-rich repeat kinase 2), including Ser910, Ser935, Ser955 and Ser973, is important for PD (Parkinson's disease) pathogenesis as several PD-linked LRRK2 mutants are dephosphorylated at these sites. LRRK2 is also dephosphorylated in cells after pharmacological inhibition of its kinase activity, which is currently proposed as a strategy for disease-modifying PD therapy. Despite this importance of LRRK2 dephosphorylation in mutant LRRK2 pathological mechanism(s) and in LRRK2's response to inhibition, the mechanism by which this occurs is unknown. Therefore we aimed to identify the phosphatase for LRRK2. Using a panel of recombinant phosphatases, we found that PP1 (protein phosphatase 1) efficiently dephosphorylates LRRK2 in vitro. PP1 activity on LRRK2 dephosphorylation was confirmed in cells using PP1 inhibition to reverse LRRK2 dephosphorylation induced by the potent LRRK2 kinase inhibitor LRRK2-IN1 as well as in R1441G mutant LRRK2. We also found that PP1 and LRRK2 can form a complex in cells. Furthermore, we observed that PP1 inhibition modulates LRRK2's cellular phenotype by reducing skein-like LRRK2-positive structures associated with dephosphorylation. In conclusion, the present study reveals PP1 as the physiological LRRK2 phosphatase, responsible for LRRK2 dephosphorylation observed in PD mutant LRRK2 and after LRRK2 kinase inhibition.
*Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, KU Leuven, Kapucijnenvoer 33, 3000 Leuven, Belgium.
Evy
E
Lobbestael
Jing
J
Zhao
Iakov N
IN
Rudenko
Aleksandra
A
Beylina
Fangye
F
Gao
Justin
J
Wetter
Monique
M
Beullens
Mathieu
M
Bollen
Mark R
MR
Cookson
Veerle
V
Baekelandt
R Jeremy
RJ
Nichols
Jean-Marc
JM
Taymans
D000818:Animals; D001570:Benzodiazepinones / Q000494:pharmacology; D002460:Cell Line; D006801:Humans; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D051379:Mice; D009154:Mutation; D010300:Parkinson Disease / Q000201:enzymology; D010766:Phosphorylation; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology
D001570:Benzodiazepinones; C582847:LRRK2-IN1; D011743:Pyrimidines; C495280:LRRK2 protein, human; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D017346:Protein Serine-Threonine Kinases; D054645:Protein Phosphatase 1