Abstract
Mutations enhancing the kinase activity of LRRK2 cause Parkinson’s disease (PD) and therapies that reduce LRRK2 kinase activity are being tested in clinical trials. Numerous rare variants of unknown clinical significance have been reported, but how the vast majority impact on LRRK2 function is unknown. Here, we investigate 98 LRRK2 variants linked to PD, including previously described pathogenic mutations, and identify 22 variants that robustly stimulate LRRK2 kinase activity. These include variants within the N-terminal non-catalytic regions [ARM (E334K, A419V), ANK (R767H), LRR (R1067Q, R1325Q)] as well as variants residing within the GTPase domain, predicted to destabilise the ROC:CORB interface [ROC (A1442P, V1447M), CORB (S1761R, L1795F)] and CORB:CORB dimer interface [CORB (R1728H/L)]. Most activating variants decrease LRRK2 biomarker site phosphorylation (pSer910/pSer935), consistent with the notion that the active kinase conformation blocks their phosphorylation. We find that the R1628P variant that has been linked to PD does not stimulate LRRK2 activity. We observe that the impact of variants on kinase activity is best evaluated by deploying a cellular assay of LRRK2-dependent Rab10 substrate phosphorylation at Thr73 compared to an in vitro kinase assay, as only a minority of activating variants enhance the activity of immunoprecipitated LRRK2, comprising the CORB (Y1699C, R1728H, S1761R) and kinase (G2019S, I2020T, T2031S) variants. Twelve variants including some that activated LRRK2, suppressed microtubule association in the presence of a Type 1 kinase inhibitor [ARM (M712V), LRR (R1320S), ROC (A1442P, K1468E, S1508R), CORA (A1589S), CORB (Y1699C, R1728H/L) and WD40 (R2143M, S2350I, G2385R)]. Our findings will aid in understanding the impact of these variants on LRRK2 kinase activation, stimulate work to reveal mechanisms by which variants impact biology, and provide rationale for variant carrier inclusion or exclusion in ongoing and future LRRK2 clinical trials.
Competing Interest Statement
The authors have declared no competing interest.
