Background:LRRK2 variants are major contributors to Parkinson's disease (PD). Many pathogenic variants increase kinase activity, underscoring the value of functional assays in nominating therapeutic targets and kinase inhibitors as potential disease-modifying therapies. Objective:To develop an interactive resource that provides functional context and ancestry-specific variant frequencies. Methods:Genotyping and short-read sequencing data were analyzed for 101,678 individuals (61,709 PD, 39,969 controls) from the Global Parkinson's Genetics Program (GP2) and integrated with clinical and in-vitro biochemical kinase activity information. Results:The LRRK2 Browser (http://gp2.org/lrrk2browser) displays ancestry-specific genetic data for 19,596 LRRK2 variants (968 exonic, 14 disease-associated) across 11 populations, and functional data for 171 variants. Clinical annotations include age, age at onset, and family history of PD. Discussion:The publicly available LRRK2 Browser represents an open-access, multi-ancestry resource to support LRRK2 variant interpretation. It aims to enhance the translational potential of genetic and functional data for precision medicine and the implementation of gene-targeted therapies in diverse populations.

Background. The VPS35 p.D620N variant causes autosomal dominant Parkinson's disease (PD) and has been shown to activate the LRRK2 kinase pathway, resulting in increased Rab substrate phosphorylation in peripheral immune cells and elevated urinary bis(monoacylglycero)phosphate (BMP) levels. Recently, a VPS35 variant of unknown significance (c.959C>T; p.A320V) was described in two late-onset sporadic PD patients. Methods. We ascertained a family from the Canary Islands in which six siblings were chronically exposed to high doses of pesticides. Three siblings developed levodopa-responsive, akinetic-rigid PD, while the other three remained unaffected. Whole-exome sequencing was performed in the three affected siblings. The frequency of the resulting candidate variant was assessed in 23,327 PD patients and 9,235 controls from four independent cohorts. Members of this pedigree and unrelated controls were assessed for LRRK2 kinase activity in monocytes and neutrophils and BMP levels in urine. Results. The three affected siblings were all heterozygous for p.A320V, whereas the three unaffected siblings did not carry the variant. In the combined PD case-control cohort, p.A320V was identified in six patients and one control. However, unlike p.D620N, heterozygous carrier status for p.A320V was not associated with increased LRRK2 kinase activity or elevated urine BMP levels. Conclusions. While VPS35 p.A320V co-segregated with PD in this family, it did not exhibit the characteristic LRRK2-associated biomarker signature observed in VPS35 p.D620N carriers. It is possible that p.A320V exerts a subtle effect on VPS35 function that was not captured by the assays performed and that chronic pesticide exposure contributed to disease penetrance in this pedigree.

Mutations in Leucine-rich repeat kinase 2 (LRRK2) and PTEN-induced kinase 1 (PINK1) are associated with familial Parkinson’s disease (PD). LRRK2 phosphorylates Rab guanosine triphosphatase (GTPases) within the Switch II domain while PINK1 directly phosphorylates Parkin and ubiquitin (Ub) and indirectly induces phosphorylation of a subset of Rab GTPases. Herein we have crossed LRRK2 [R1441C] mutant knock-in mice with PINK1 knock-out (KO) mice and report that loss of PINK1 does not impact endogenous LRRK2-mediated Rab phosphorylation nor do we see significant effect of mutant LRRK2 on PINK1-mediated Rab and Ub phosphorylation. In addition, we observe that a pool of the Rab-specific, protein phosphatase family member 1H phosphatase, is transcriptionally up-regulated and recruited to damaged mitochondria, independent of PINK1 or LRRK2 activity. Parallel signaling of LRRK2 and PINK1 pathways is supported by assessment of motor behavioral studies that show no evidence of genetic interaction in crossed mouse lines. Previously we showed loss of cilia in LRRK2 R1441C mice and herein we show that PINK1 KO mice exhibit a ciliogenesis defect in striatal cholinergic interneurons and astrocytes that interferes with Hedgehog induction of glial derived-neurotrophic factor transcription. This is not exacerbated in double-mutant LRRK2 and PINK1 mice. Overall, our analysis indicates that LRRK2 activation and/or loss of PINK1 function along parallel pathways to impair ciliogenesis, suggesting a convergent mechanism toward PD. Our data suggest that reversal of defects downstream of ciliogenesis offers a common therapeutic strategy for LRRK2 or PINK1 PD patients, whereas LRRK2 inhibitors that are currently in clinical trials are unlikely to benefit PINK1 PD patients.

Background Gain-of-kinase-function variants in LRRK2 are a leading cause of monogenic Parkinson's disease (PD). Objectives We tested the functional impact of a novel LRRK2 variant p.V1447L identified in a young-onset PD patient in vivo in peripheral blood, as well as in a robust cellular assay, alongside other variants in close proximity to V1447. Methods We measured LRRK2-dependent Rab10 phosphorylation in neutrophils and monocytes of a LRRK2 p.V1447L carrier with PD. We performed structural mapping and evaluated the potential impact of other LRRK2 variants at and around LRRK2 V1447. Results LRRK2 p.V1447L strongly increases LRRK2 kinase activity. We identified additional variants in the LRRK2 ROC:CORB interface with critical impact on kinase activity and demonstrated that different substitutions at the same residue can have opposing effects. Conclusions We recommend reclassifying LRRK2 p.V1447L from variant of uncertain significance to likely pathogenic. Our study expands the range of putative loss-of-kinase function variants to LRRK2 missense variants. (c) 2025 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

The end-stage pathology of Parkinson's disease (PD) involves the loss of dopamine-producing neurons in the substantia nigra pars compacta (SNc). However, synaptic deregulation of these neurons begins much earlier. Understanding the mechanisms behind synaptic deficits is crucial for early therapeutic intervention, yet these remain largely unknown. In the SNc, different dopamine neuron subtypes show varying susceptibility patterns to PD, complicating our understanding. This study uses intersectional genetic mouse models to uncover synaptic perturbations in vulnerable dopamine neurons, focusing on the LRRK2 kinase, a protein closely linked to PD. Through a combination of immunofluorescence and advanced proximity labeling methods, we found higher LRRK2 expression in the most vulnerable dopamine neuron subclusters. High-resolution imaging revealed that pathogenic LRRK2 disrupts release sites in vulnerable dopamine axons, leading to decreased in vivo evoked striatal dopamine release in mice with LRRK2 mutations. Proteomic and biochemical analyses indicate that mutant LRRK2 increases the phosphorylation of RAB3 proteins, reducing their interactions with RIM1/2 effector proteins and impacting their synaptic functions. Overall, this research highlights the cell-autonomous dysfunctions caused by mutant LRRK2 in the neurons that are primarily affected by the disease. It also provides a framework for therapeutic strategies for early nigrostriatal synaptic deficits in PD.

Parkinson's disease is associated with activating mutations in the gene encoding leucine-rich repeat kinase 2 (LRRK2), which suppresses primary cilia formation in cholinergic and parvalbumin interneurons and astrocytes in the striatum. As a result, there is a decrease in the production of neuroprotective glial cell line-derived neurotrophic factor (GDNF) and neurturin (NRTN), which normally support the viability of dopaminergic neurons. MLi-2 is a brain-penetrant, selective, and now experimental inhibitor of LRRK2. Here, we found that dietary administration of MLi-2 to young LRRK2-mutant mice for 3 months restored primary cilia formation and Hedgehog signaling in both cholinergic and parvalbumin interneurons and astrocytes. The treatment also restored the Hedgehog-responsive expression of Gdnf and Nrtn in the neurons. Cilia were also restored on cholinergic neurons of the pedunculopontine nucleus, where their loss correlates with severity of motor impairment in patients. Furthermore, MLi-2 increased the density of fine striatal dopaminergic processes and decreased the amount of stress-associated Sonic Hedgehog RNA expression in nigral dopaminergic neurons. Thus, pathogenic LRRK2-driven cilia loss is reversible in postmitotic neurons and astrocytes, which suggests that early administration of specific LRRK2 inhibitors may therapeutically benefit patients.

Dissecting biological pathways highlighted by Mendelian gene discovery has provided critical insights into the pathogenesis of Parkinson's disease (PD) and neurodegeneration. This approach ultimately catalyzes the identification of potential biomarkers and therapeutic targets. Here, we identify PSMF1 as a novel gene implicated in parkinsonism and childhood neurodegeneration. We find that biallelic PSMF1 missense and loss-of-function variants co-segregate with phenotypes from early-onset PD to perinatal lethality with neurological manifestations across 17 pedigrees with 24 affected subjects, showing clear genotype-phenotype correlation. PSMF1 encodes the proteasome regulator PSMF1/PI31, a highly conserved, ubiquitously expressed partner of the 20S proteasome and neurodegeneration-associated F-box-O 7 and valosin-containing proteins. We demonstrate that PSMF1 variants impair mitochondrial membrane potential, dynamics and mitophagy, and may affect proteasomal abundance and assembly in patient-derived fibroblasts. Furthermore, Drosophila and mouse models of PSMF1 loss of function exhibit age-dependent motor impairment, as well as brain-wide mitochondrial membrane depolarization and dopaminergic neurodegeneration in aged flies, and diffuse gliosis in mice. Collectively, our findings unequivocally link defective PSMF1 to early-onset parkinsonism and neurodegeneration, and suggest proteasomal and mitochondrial dysfunction as mechanistic contributors.

While mRNA splicing dysregulation is a well-established contributor to neurodegeneration in disorders such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), its role in Parkinson’s disease (PD) remains underexplored. Here, we analyse transcriptomic data from >500 post-mortem human brain samples from individuals with and without PD to show that splicing alterations are frequently detected. Differentially spliced genes were significantly more enriched for those causally-implicated in both PD and ALS than genes that were differentially expressed. Furthermore, we observed a strong association between these splicing alterations and dysfunction of the RNA-binding protein (RBP), TAR DNA-binding protein 43 (TDP-43). Strikingly, genes and exon junctions affected by TDP-43 knockdown overlapped significantly with those dysregulated across brain regions in PD. In brains from individuals with the LRRK2 c.6055G>A (p.G2019S) mutation, the most common genetic cause of PD, we also observed significant enrichment of TDP-43-dependent splicing changes. This finding was corroborated in human pluripotent stem cell-derived midbrain dopaminergic neurons and a LRRK2 p.G2019S knock-in mouse model, where reduced nuclear TDP-43 levels evidenced the well-recognised loss-of-function mechanism contributing to splicing dysregulation. By leveraging our RNA-based analyses we predicted TDP-43-dependent novel peptide sequences and validated their existence within human LRRK2 mutation mDNs, while also demonstrating an overall loss of protein and mRNA expression in mis-spliced genes. Collectively, our findings reveal that PD is marked by extensive splicing dysregulation dependent on TDP-43, making TDP-43 a promising new therapeutic target in PD. ### Competing Interest Statement The authors have declared no competing interest. Aligning Science Across Parkinson's, https://ror.org/03zj4c476, ASAP-000478, ASAP-000509, ASAP-000486

The past 10 years have seen tremendous progress in our understanding of leucine-rich repeat kinase 2 (LRRK2) and how mutations activate the kinase and trigger downstream pathology, contributing to Parkinson's disease. A breakthrough came from the identification of key LRRK2 substrates-a subset of small guanosine triphosphatases (GTPases) called Rab proteins. Cryoelectron microscopy has revealed structures of LRRK2 and showed how inhibitors engage and inhibit the kinase. Biochemical experiments have revealed how LRRK2 is recruited to membranes to phosphorylate Rab substrates. LRRK2 activation during lysosomal stress triggers Rab phosphorylation, altering the repertoire of Rab-binding partners. Resulting phospho-Rab-effector complexes have prominent effects in specific cell types, disrupting primary cilia and impairing Hedgehog signaling-effects that can be reversed by LRRK2 inhibitors. This disruption in Hedgehog signaling represents a convergence point linking genetic and idiopathic forms of Parkinson's. Together, these findings support the therapeutic potential of LRRK2 inhibitors in Parkinson's disease.

Leucine-rich repeat kinase 2 (LRRK2) inhibition is a promising disease-modifying therapy for LRRK2-associated Parkinson's disease (L2PD) and idiopathic PD. However, pharmacodynamic readouts and progression biomarkers for clinical trials aiming for disease modification are insufficient, given that no endogenous marker reflecting enhanced kinase activity of the most common LRRK2 G2019S mutation has yet been reported in L2PD patients.Using phospho-/proteomic analyses, we assessed the impact of LRRK2-activating mutations in peripheral blood mononuclear cells from an LRRK2 clinical cohort from Spain (n = 174). The study groups encompassed G2019S L2PD patients (n = 37), non-manifesting LRRK2 mutation carriers of G2019S (here termed G2019S L2NMCs) (n = 27), R1441G L2PD patients (n = 14), R1441G L2NMCs (n = 11), idiopathic PD patients (n = 40) and healthy controls (n = 45).We identified 207 differentially regulated proteins in G2019S L2PD compared with controls (39 upregulated and 168 downregulated) and 67 in G2019S L2NMCs (10 upregulated and 57 downregulated). G2019S downregulated proteins affected the endolysosomal pathway, proteostasis and mitochondria, e.g. ATIC, RAB9A or LAMP1. At the phospho-proteome level, we observed increases in endogenous phosphorylation levels of pSer106 RAB12 in G2019S carriers, which were validated by immunoblotting after 1 year of follow-up (n = 48). Freshly collected peripheral blood mononuclear cells from three G2019S L2PD, one R1441G L2PD, one idiopathic PD and five controls (n = 10) showed strong diminishment of pSer106 RAB12 phosphorylation levels after in vitro administration of the MLi-2 LRRK2 inhibitor. Using machine learning, we identified an 18-feature G2019S phospho-/protein signature discriminating G2019S L2PD, L2NMCs and controls with 96% accuracy that was correlated with disease severity, i.e. UPDRS-III motor scoring.Using easily accessible peripheral blood mononuclear cells from a LRRK2 clinical cohort, we identified elevated levels of pSer106 RAB12 as an endogenous biomarker of G2019S carriers. Our data suggest that monitoring pSer106 RAB12 phosphorylation could be a relevant biomarker for tracking LRRK2 activation, particularly in G2019S carriers. Future work might determine whether pSer106 RAB12 could help with patient enrichment and monitoring drug efficacy in LRRK2 clinical trials. The LRRK2 activating mutation G2019S is the most frequent genetic cause of Parkinson's disease. Through phospho-proteome analysis of blood, Cort & eacute;s et al. identify elevated phospho-RAB12 levels as an endogenous biomarker of G2019S mutation carriers, with potential utility in clinical trials.

WNK family kinases are regulated by osmotic stress and control ion homeostasis by activating SPAK and OXSR1 kinases. Using a proximity labeling approach, we found that osmotic stress promotes the association of WNK1 with the NRBP1 pseudokinase and TSC22D2/4 adaptor proteins, results that are confirmed by immunoprecipitation, mass spectrometry, and immunoblotting studies. NRBP1 pseudokinase is closely related to WNK isoforms and contains a RΦ-motif–binding conserved C-terminal (CCT) domain, like the CCT domains in WNKs, SPAK, and OXSR1. Knockdown or knockout of NRBP1 markedly inhibited basal as well as sorbitol-induced activation of WNK1 and downstream components. We demonstrate that recombinant NRBP1 can directly induce the activation of WNK4 in vitro. AlphaFold-3 modeling predicts that WNK1, SPAK, NRBP1, and TSC22D4 form a complex, in which two TSC22D4 RΦ-motifs interact with the CCTL1 domain of WNK1 and the CCT domain of NRBP1. Our data indicate that NRBP1 and likely its close homolog NRBP2 function as an upstream activator of the WNK pathway.

LRRK2-related Parkinson's disease (LRRK2-PD) is the most frequent form of monogenic PD worldwide, with important therapeutic opportunities, exemplified by the advancement in LRRK2 kinase inhibition studies/trials. However, many LRRK2 variants, especially those found in underrepresented populations, remain classified as variants of uncertain significance (VUS). Leveraging on Malaysian, Singaporean, and mainland Chinese PD datasets (n = 4901), we describe 12 Chinese-ancestry patients harboring the LRRK2 p.Arg1067Gln variant, more than doubling the number of previously reported cases (total n = 23, 87% East Asian, mean age of onset: 53.9 years). We determine that this variant is enriched in East Asian PD patients compared to population controls (OR = 8.0, 95% CI: 3.0-20.9), and provide supportive data for its co-segregation with PD, albeit with incomplete penetrance. Utilizing established experimental workflows, this variant showed increased LRRK2 kinase activity, by similar to 2-fold compared to wildtype and higher than the p.Gly2019Ser variant. Taken together, p.Arg1067Gln should be reclassified from a VUS to pathogenic for causing LRRK2-PD.

Lysosomes are implicated in a wide spectrum of human diseases, including monogenic lysosomal storage disorders (LSDs), age-associated neurodegeneration, and cancer. Profiling lysosomal content using tag-based lysosomal immunoprecipitation (LysoTagIP) in cell and animal models has substantially moved the field forward, but studying lysosomal dysfunction in patients remains challenging. Here, we report the development of the 'tagless LysoIP' method, designed to enable the rapid enrichment of lysosomes, via immunoprecipitation, using the endogenous integral lysosomal membrane protein TMEM192, directly from clinical samples and human cell lines (e.g., induced pluripotent stem cell-derived neurons). Isolated lysosomes were intact and suitable for subsequent multimodal omics analyses. To validate our approach, we applied the tagless LysoIP to enrich lysosomes from peripheral blood mononuclear cells derived from fresh blood of healthy donors and patients with CLN3 disease, an autosomal recessive neurodegenerative LSD. Metabolic profiling of isolated lysosomes revealed massive accumulation of glycerophosphodiesters (GPDs) in patients' lysosomes. Interestingly, a patient with a milder phenotype and genotype displayed lower accumulation of lysosomal GPDs, consistent with their potential role as disease biomarkers. Altogether, the tagless LysoIP provides a framework to study native lysosomes from patient samples, identify disease biomarkers, and discover human-relevant disease mechanisms.

A GGGGCC repeat expansion in C9orf72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). The repeat expansion is translated into five different dipeptide repeat proteins: polyGA, polyGP, polyGR, polyAP and polyPR. To investigate the effect of polyGA, which is the most abundant dipeptide repeat protein in patient brains, we used CRISPR/Cas9 to insert 400 codon-optimized polyGA repeats immediately downstream of the mouse C9orf72 start codon. This generated (GA)400 knock-in mice driven by the endogenous mouse C9orf72 promoter, coupled with heterozygous C9orf72 reduction. (GA)400 mice develop subtle pathology including mild motor dysfunction characterized by impaired rotarod performance. Quantitative proteomics revealed polyGA expression caused protein alterations in the spinal cord, including changes in previously identified polyGA interactors. Our findings show that (GA)400 mice are a complementary in vivo model to better understand C9ALS/FTD pathology and determine the specific role of single DPRs in disease. ### Competing Interest Statement The authors have declared no competing interest.

Introduction: WNK kinases phosphorylate and activate SPAK/OSR1, which phosphorylate and activate the Na + -Cl - cotransporter (NCC) in the distal convoluted tubule (DCT) of the kidney. WNK4 participates in NCC regulation by plasma K + . Under elevated plasma K + , WNK4 activity is inhibited by Cl - binding. Under low plasma K + , [Cl - ] i decreases, relieving Cl - inhibition of WNK4. Additionally, WNK biomolecular condensates (WNK bodies) form in the DCT under low K + stress. In Familial hyperkalemic hypertension (FHHt), caused by NCC overactivation, WNK condensate formation in the DCT occurs due to mutations that impair WNK degradation and lead to WNK overexpression without directly increasing kinase activity. WNK condensates are also formed in cells that undergo osmotic stress. Whether WNK condensate formation drives kinase activation or merely correlates with catalytic activity, and the mechanism by which this occurs, remains unknown. We hypothesize that WNK condensates activate WNK kinases by forming Cl - -reduced microdomains, underpinning the pathophysiology of FHHt. Methods: Constructs of wild-type or chloride-insensitive WNK1, WNK3, and WNK4 fused to mCherry and Cry2clust enabled optogenetic condensate induction. The WNK1 kinase domain was fused to the intrinsically disordered region (IDR) of FUS, TDP-43, or DDX3X, and Cry2clust. A WNK4-ClopHensorN fusion for ratiometric Cl - measurement within WNK bodies was created. A novel pharmacogenetic model of phase separation was generated by fusing WNK4 to a tamoxifen-sensitive estrogen receptor ligand-binding domain. FHHt mice (homozygous KLHL3 R528H mutants) were subjected to a 5% K + diet to evaluate NCC phosphorylation, WNK body assembly, and K + homeostasis. Mice lacking KS-WNK1 in the KLHL3 R528H background were used to assess WNK body formation. Western Blot of total and phosphorylated forms of the proteins of interest was performed. Live-cell fluorescence imaging was done to assess the induction of biomolecular condensation and for Cl - measurement with ClopHensorN. Immunofluorescent staining of WNK body components was carried out along with that of NCC to mark DCT cells. Plasma electrolytes were measured using an iStat device. Statistical analyses included unpaired t-tests for two-group comparisons and two-way ANOVA for multiple groups, with an alpha value of 0.05. Results: Optogenetic and pharmacologic induction of WNK oligomerization promoted condensation and activated full-length WT WNK kinases (measured by SPAK phosphorylation), but not Cl - -insensitive WNK kinases, suggesting a Cl - -sensing-dependent activation mechanism. Autophosphorylation in the T-loop (S382) of WNK1 in condensates occurred with its native IDR, but not with FUS, TDP-43, or DDX3X fusions, suggesting that properties conferred by the WNK IDR are essential for condensation-induced activation. [Cl - ] in WNK bodies, measured with ClopHensorN, approximated R0Cl (fluorescence intensity ratio in 0 mM [Cl - ]). FHHt mice displayed WNK bodies and were unable to downregulate pNCC during a high-K + diet, even with severe hyperkalemia (mean plasma [K + ]: 8.8 mM vs. 5.2 mM in WT; p<0.0001 ). Surprisingly, KLHL3 R528H mice lacking KS-WNK1 still showed WNK4-positive condensates in their DCT, indicating that WNK4 overexpression suffices for condensate assembly in FHHt. Conclusion: WNK body formation drives kinase activation and represents a mechanism that regulates NCC and, very likely, K + homeostasis. Our experiments suggest that condensation-induced activation may be due to the formation of Cl - -reduced microdomains, and could be the cause of the FHHt phenotype by preventing kinase inactivation under the high intracellular Cl - conditions expected in this disease. CONAHCyT grant 737116 to HCC NIH R01 grant DK51496 to DE and GG Kidneycure TIG to MCB This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

Lysosomes are implicated in a wide spectrum of human diseases including monogenic lysosomal storage disorders (LSDs), age-associated neurodegeneration and cancer. Profiling lysosomal content using tag-based lysosomal immunoprecipitation (LysoTagIP) in cell and animal models allowed major discoveries in the field, however studying lysosomal dysfunction in human patients remains challenging. Here, we report the development of the “tagless LysoIP method” to enable rapid enrichment of lysosomes, via immunoprecipitation, using the endogenous integral lysosomal membrane protein TMEM192, directly from clinical samples and human cell lines (e.g. induced Pluripotent Stem Cell (iPSCs) derived neurons). Isolated lysosomes are intact and suitable for subsequent multimodal omics analyses. To validate our approach, we employed the tagless LysoIP to enrich lysosomes from peripheral blood mononuclear cells (PBMCs) derived from fresh blood from patients with CLN3 disease, a neurodegenerative LSD. Metabolic profiling of isolated lysosomes showed massive accumulation of glycerophosphodiesters (GPDs) in patients’ lysosomes. Interestingly, a patient with a milder phenotype and genotype displayed lower accumulation of lysosomal GPDs, consistent with their potential role as disease biomarkers. Altogether, the tagless LysoIP provides a framework to study native lysosomes from patient samples, identify novel biomarkers and discover human-relevant disease mechanisms.### Competing Interest StatementThe authors have declared no competing interest.

Activating LRRK2 mutations cause Parkinson's disease. Previously, we showed that cholinergic interneurons and astrocytes but not medium spiny neurons of the dorsal striatum lose primary cilia in LRRK2 mutant mice. Single nucleus RNA sequencing shows that cilia loss in cholinergic interneurons correlates with higher LRRK2 expression and decreased glial derived neurotrophic factor transcription. Nevertheless, much higher LRRK2 expression is seen in medium spiny neurons that have normal cilia in mice and humans. In parallel with decreased striatal dopaminergic neurite density, LRRK2 G2019S neurons show increased autism-linked CNTN5 adhesion protein expression; glial cells show significant loss of ferritin heavy chain. Human striatal tissue from LRRK2 pathway mutation carriers and idiopathic Parkinson's disease show similar cilia loss in cholinergic interneurons and astrocytes and overall loss of such neurons. These data strongly suggest that loss of cilia in specific striatal cell types decreases neuroprotection for dopamine neurons in mice and human Parkinson's disease.

Activating leucine-rich repeat kinase 2 (LRRK2) mutations cause Parkinson’s and phosphorylation of Rab10 by pathogenic LRRK2 blocks primary ciliogenesis in cultured cells. In the mouse brain, LRRK2 blockade of primary cilia is highly cell type specific: For example, cholinergic interneurons and astrocytes but not medium spiny neurons of the dorsal striatum lose primary cilia in LRRK2-pathway mutant mice. We show here that the cell type specificity of LRRK2-mediated cilia loss is also seen in human postmortem striatum from patients with LRRK2 pathway mutations and idiopathic Parkinson’s. Single nucleus RNA sequencing shows that cilia loss in mouse cholinergic interneurons is accompanied by decreased glial-derived neurotrophic factor transcription, decreasing neuroprotection for dopamine neurons. Nevertheless, LRRK2 expression differences cannot explain the unique vulnerability of cholinergic neurons to LRRK2 kinase as much higher LRRK2 expression is seen in medium spiny neurons that have normal cilia. In parallel with decreased striatal dopaminergic neurite density, LRRK2 G2019S neurons show increased autism-linked CNTN5 adhesion protein expression; glial cells show significant loss of ferritin heavy chain. These data strongly suggest that loss of cilia in specific striatal cell types decreases neuroprotection for dopamine neurons in mice and human Parkinson’s.