Biology subjects

Taymans, J.-M.

Publications and source records attributed to Taymans, J.-M..

6 recordsLinked to original sources

bioRxiv2026

The phosphorylation status of LRRK2 at the S910/S935 cluster determines its sensitivity to activation by RAB29

Leucine-Rich Repeat Kinase 2 (LRRK2) is a signaling molecule involved in Parkinsons disease pathomechanisms. In disease, the LRRK2 protein displays both a toxic gain of kinase function and a loss of phosphorylation at heterophosphosites found in an extended loop of the LRR domain. RAB GTPases, such as RAB29, have been identified as upstream activators of LRRK2. Indeed, co-expression of LRRK2 with RAB29 induces a hyperactivation of LRRK2 kinase activity, however the role of the LRRK2 heterologous phosphorylation status in its activation remains unknown. Here, our aim was to determine the role of LRRK2 heterologous phosphorylation on its activation by RAB29. Using single and compound phosphodead or phosphomimetic mutants of LRRK2 we show differential sensitivity of LRRK2 phosphomutants to activation by RAB29, with phosphodead mutants being more susceptible to be activated than phosphomimetic mutants. Interestingly, we find that the single phosphodead S910A LRRK2 mutant displays an activation of LRRK2 kinase activity similar to that observed for the compound phosphodead 6xS>A LRRK2 mutant (S860A/S910A/S935A/S955A/S973A/S976A). Time-course analysis revealed that phosphodead mutants displayed higher but also faster activation by RAB29. In addition, both physical interaction between LRRK2 and RAB29 as well as RAB29-induced recruitment of LRRK2 to the trans-Golgi network (TGN) was enhanced by phosphodead compared to phosphomimetic mutants. To confirm effects on native LRRK2, we tested a panel of ten nanobodies targeting LRRK2 that stabilized LRRK2 phosphorylation at varying levels. Nanobodies stabilizing LRRK2 at low S935 phosphorylation levels showed enhanced RAB29-induced activation compared to nanobodies not affecting pS935 LRRK2. Finally, we tested whether LRRK2 heterologous phosphorylation could affect centrosome cohesion deficits, a phenotype that has been linked to LRRK2 hyperactivation, and found that both the phosphodead LRRK2 as well as a nanobody stabilizing dephosphorylated LRRK2 enhanced the centrosome cohesion deficit. Our findings indicate that hyperactivability of LRRK2 is directly related to its heterologous phosphorylation status, with dephosphorylation leading to strong hyperactivation of LRRK2 by upstream activating RABs, and phosphorylated LRRK2 showing the opposite. This implies that strategies favoring LRRK2 phosphorylation will have therapeutic benefit.

neuroscience↗

bioRxiv2025

Multi-cohort, cross-species urinary proteomics reveals signatures of LRRK2 dysfunction in Parkinsons disease

Pathogenic mutations in Leucine-rich repeat kinase 2 (LRRK2) are the predominant genetic cause of Parkinsons disease (PD) and often increase kinase activity, making LRRK2 inhibitors promising treatment options. Although LRRK2 kinase inhibitors are advancing clinically, non-invasive readouts of LRRK2-linked pathway modulation remain limited. Profiling urinary proteomes from 1,215 individuals across three cohorts and integrating whole-genome sequencing from >500 participants to map genotype-proteome associations, we identified 177 urinary proteins associated with pathogenic LRRK2, enriched for lysosomal/glycosphingolipid, immune, and membrane-trafficking pathways. Machine learning narrowed the features to a cohort-agnostic 30-protein panel that classified G2019S carriers with a mean ROC AUC of 0.91 across independent tests. To evaluate translation, we performed multi-organ and urinary proteomics in rat gain- and loss-of-function models (BAC-LRRK2G2019S and Lrrk2KO) and after Lrrk2 inhibition (MLi-2 and PF-475), revealing tissue-specific responses--strongest in kidney--and cross-species overlap, including 24 brain proteins detectable in human urine. Rat-derived perturbations predicted LRRK2 mutation status in patients (AUC 0.75) and reversed with Lrrk2 inhibition, supporting their pharmacodynamic utility. Together, our findings establish urine as a scalable, non-invasive matrix that captures systemic and brain-relevant consequences of LRRK2 dysfunction and nominate candidate pharmacodynamic markers set to support LRRK2-directed trials.

systems biology↗

bioRxiv2025

A regulatory loop regulating signaling between LRRK2 and PP2A in cellular models of Parkinsons disease

Mutations in Leucine-rich repeat kinase 2 (LRRK2) are the most frequent cause of late-onset familial and idiopathic Parkinsons disease (PD), known to date. Importantly, recent data from post-mortem tissue as well as biomarker studies suggest that independent of mutations, increased kinase activity of LRRK2 plays an essential role in idiopathic PD pathogenesis. Despite extensive research on LRRK2, its activation mechanism(s) and how the various mutations result in increased kinase activity and neuronal death is still not completely understood. Accumulating evidence points to LRRK2 phospho-regulation, both auto-phosphorylation and phosphorylation by other kinases, as one potential molecular trigger of its activation. LRRK2 activation and localization is regulated by phosphatases such as Protein phosphatase 1 (PP1) and Protein phosphatase 2A (PP2A), however the exact mechanism of this phospho-regulation is not known. Our data reveal that in vitro PP2A dephosphorylates sites within the RocCOR-GTPase domain of LRRK2 and as a result de-stabilizes LRRK2 dimers, with consequent reduction of its kinase activity. Strikingly, our data further highlight that LRRK2 in turn phosphorylates the catalytic subunit of the PP2A holoenzyme PPP2CA at its critical residue T304. Furthermore, LRRK2-mediated phosphorylation of PP2CA T304 alters the methylation of the C-terminus, which is crucial for both holoenzyme formation and catalytic activity. Importantly, expression of WT-PPP2CA protects from LRRK2-G2019S induced neuronal cell death, while PPP2CA-T304 mutants fail to do so, suggesting that impaired PP2A holoenzyme formation might be detrimental for LRRK2-PD.

neuroscience↗

bioRxiv2024

PAK6 rescues pathogenic LRRK2-mediated ciliogenesis and centrosomal cohesion defects in a mutation-specific manner

P21 activated kinase 6 (PAK6) is a serine-threonine kinase with physiological expression enriched in the brain and overexpressed in a number of human tumors. While the role of PAK6 in cancer cells has been extensively investigated, the physiological function of the kinase in the context of brain cells is poorly understood. Our previous work uncovered a link between PAK6 and the Parkinsons disease (PD)-associated kinase LRRK2, with PAK6 controlling LRRK2 activity and subcellular localization via phosphorylation of 14-3-3 proteins. Here, to gain more insights into PAK6 physiological function, we performed protein-protein interaction arrays and identified a subgroup of PAK6 binders related to ciliogenesis. We confirmed that endogenous PAK6 localizes at both the centrosome and the cilium, and positively regulates ciliogenesis not only in tumor cells but also in neurons and astrocytes. Strikingly, PAK6 rescues ciliogenesis and centrosomal cohesion defects associated with the G2019S but not the R1441C LRRK2 PD mutation. Since PAK6 binds LRRK2 via its GTPase/Roc-COR domain and the R1441C mutation is located in the Roc domain, we used microscale thermophoresis and AlphaFold2-based computational analysis to demonstrate that PD mutations in LRRK2 affecting the Roc-COR structure substantially decrease PAK6 affinity, providing a rationale for the differential protective effect of PAK6 toward the distinct forms of mutant LRRK2. Altogether, our study discloses a novel role of PAK6 in ciliogenesis and points to PAK6 as the first LRRK2 modifier with PD mutation-specificity.

neuroscience↗

bioRxiv2023

A potential patient stratification biomarker for Parkinson's disease based on LRRK2 kinase-mediated centrosomal alterations in peripheral blood-derived cells

Parkinsos disease (PD) is a common neurodegenerative movement disorder and leucine-rich repeat kinase 2 (LRRK2) is a promising therapeutic target for disease intervention. However, the ability to stratify patients who will benefit from such treatment modalities based on shared etiology is critical for the success of disease-modifying therapies. Ciliary and centrosomal alterations are commonly associated with pathogenic LRRK2 kinase activity and can be detected in many cell types. We previously found centrosomal deficits in immortalized lymphocytes from G2019S-LRRK2 PD patients. Here, to investigate whether such deficits may serve as a potential blood biomarker for PD which is susceptible to LRKK2 inhibitor treatment, we characterized patient-derived cells from distinct PD cohorts. We report centrosomal alterations in peripheral cells from a subset of early-stage idiopathic PD patients which is mitigated by LRRK2 kinase inhibition, supporting a role for aberrant LRRK2 activity in idiopathic PD. Centrosomal defects are detected in R1441G-LRRK2 and G2019S-LRRK2 PD patients and in non-manifesting LRRK2 mutation carriers, indicating that they acumulate prior to a clinical PD diagnosis. They are present in immortalized cells as well as in primary lymphocytes from peripheral blood. These findings indicate that analysis of centrosomal defects as a blood-based patient stratification biomarker may help nominate PD patients who will benefit from LRRK2-related therapeutics. One-sentence summaryPeripheral blood-derived cells can be employed to stratify Parkinsos disease patients most likely to respond to LRRK2-related therapeutics.

neuroscience↗

bioRxiv2021

Functional analyses of two novel LRRK2 pathogenic variants in familial Parkinson's disease

BackgroundPathogenic variants in the LRRK2 gene are a common monogenic cause of Parkinsons disease. However, only seven variants have been confirmed to be pathogenic. ObjectivesWe identified two novel LRRK2 variants (H230R and A1440P) and performed functional testing. MethodsWe transiently expressed wildtype, the two new variants, or two known pathogenic mutants (G2019S and R1441G), in HEK-293T cells, with or without LRRK2 kinase inhibitor treatment. We characterized the phosphorylation and kinase activity of the mutants by western blotting. Thermal shift assays were performed to determine the folding and stability of the LRRK2 proteins. ResultsThe two variants were found in two large families and segregate with the disease. They display altered LRRK2 phosphorylation and kinase activity. ConclusionsWe identified two novel LRRK2 variants which segregate with the disease. The results of functional testing lead us to propose these two variants as novel causative mutations for familial Parkinsons disease.

genetics↗