Cellular processes associated with LRRK2 function and dysfunction.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) encoding gene are the most common cause of monogenic Parkinson's Disease (PD). The identification of LRRK2 polymorphisms associated with increased risk for sporadic PD, as well as the observation that LRRK2-PD has an almost indistinguishable pathological phenotype from the sporadic form of disease, suggested LRRK2 as the culprit to provide understanding for both familial and sporadic PD cases. LRRK2 is a large protein with both GTPase and kinase functions. Mutations segregating with PD reside within the enzymatic core of LRRK2, suggesting the modification of its activity greatly impacts disease onset and progression. Although progress has been gained since its discovery in 2004, there is still much to be understood regarding LRRK2's physiological and neurotoxic properties. Unsurprisingly, given the presence of multiple enzymatic domains, LRRK2 has been associated with a diverse set of cellular functions and signalling pathways including mitochondrial function, vesicle trafficking together with endocytosis, retromer complex modulation and autophagy. This review will discuss the state of current knowledge for the role of LRRK2 in health and disease with discussion of potential substrates of phosphorylation and functional partners with particular emphasis on signalling mechanisms. As well, the use of immune cells in LRRK2 research and the role of oxidative stress as a regulator of LRRK2 activity and cellular function shall also be discussed. This article is protected by copyright. All rights reserved

WHOPPA Enables Parallel Assessment of Leucine-Rich Repeat Kinase 2 and Glucocerebrosidase Enzymatic Activity in Parkinson's Disease Monocytes

Both leucine-rich repeat kinase 2 (LRRK2) and glucocerebrosidase (GCase) are promising targets for the treatment of Parkinson's disease (PD). Evidence suggests that both proteins are involved in biological pathways involving the lysosome. However, studies to date have largely investigated the enzymes in isolation and any relationship between LRRK2 and GCase remains unclear. Both enzymes are highly expressed in peripheral blood monocytes and have been implicated in immune function and inflammation. To facilitate the standardized measurement of these readouts in large cohorts of samples collected from persons with PD across the globe, we developed and optimized a sample collection and processing protocol with parallel flow cytometry assays. Assay parameters were first optimized using healthy control peripheral blood mononuclear cells (PBMCs), and then LRRK2 and GCase activities were measured in immune cells from persons with idiopathic PD (iPD). We tested the ability of this protocol to deliver similar results across institutes across the globe, and named this protocol the Wallings-Hughes Optimized Protocol for PBMC Assessment (WHOPPA). In the application of this protocol, we found increased LRRK2 levels and stimulation-dependent enzymatic activity, and decreased GBA index in classical iPD monocytes, as well as increased cytokine release in PD PBMCs. WHOPPA also demonstrated a strong positive correlation between LRRK2 levels, pRab10 and HLA-DR in classical monocytes from subjects with iPD. These data support a role for the global use of WHOPPA and expression levels of these two PD-associated proteins in immune responses, and provide a robust assay to determine if LRRK2 and GCase activities in monocytes have potential utility as reliable and reproducible biomarkers of disease in larger cohorts of subjects with PD.Copyright © 2022 Wallings, Hughes, Staley, Simon, McFarland, Alcalay, Garrido, Martí, Sarró, Dzamko and Tansey

The BAT-Swift Science Software

The BAT instrument tells the Swift satellite where to point to make immediate follow-up observations of GRBs. The science software on board must efficiently process gamma-ray events coming in at up to 34 kHz, identify rate increases that could be due to GRBs while disregarding those from known sources, and produce images to accurately and rapidly locate new Gamma-ray sources.Comment: 4 pages, no figures, to appear in Santa Fe proceedings "Gamma-Ray Bursts: 30 Years of Discovery", Fenimore and Galassi (eds), AIP, 200

Progranulin Loss Results in Sex-Dependent Dysregulation of the Peripheral and Central Immune System

INTRODUCTION: Progranulin (PGRN) is a secreted glycoprotein, the expression of which is linked to several neurodegenerative diseases. Although its specific function is still unclear, several studies have linked it with lysosomal functions and immune system regulation. Here, we have explored the role of PGRN in peripheral and central immune system homeostasis by investigating the consequences of PGRN deficiency on adaptive and innate immune cell populations. METHODS: First, we used gene co-expression network analysis of published data to test the hypothesis that RESULTS: Male PGRN KO mice exhibited a lower abundance of microglial cells with higher MHC-II expression, increased CD44 expression on monocytes in the brain, and more CNS-associated CD8 DISCUSSION: Our data suggest that PGRN and GPNMB jointly regulate the peripheral and the central immune system in a sex-specific manner; thus, understanding their associated mechanisms could pave the way for developing new neuroprotective strategies to modulate central and peripheral inflammation to lower risk for neurodegenerative diseases and possibly delay or halt progression

Gene therapy restores dopamine transporter expression and ameliorates pathology in iPSC and mouse models of infantile parkinsonism

Most inherited neurodegenerative disorders are incurable, and often only palliative treatment is available. Precision medicine has great potential to address this unmet clinical need. We explored this paradigm in dopamine transporter deficiency syndrome (DTDS), caused by biallelic loss-of-function mutations in SLC6A3, encoding the dopamine transporter (DAT). Patients present with early infantile hyperkinesia, severe progressive childhood parkinsonism, and raised cerebrospinal fluid dopamine metabolites. The absence of effective treatments and relentless disease course frequently leads to death in childhood. Using patient-derived induced pluripotent stem cells (iPSCs), we generated a midbrain dopaminergic (mDA) neuron model of DTDS that exhibited marked impairment of DAT activity, apoptotic neurodegeneration associated with TNFα-mediated inflammation, and dopamine toxicity. Partial restoration of DAT activity by the pharmacochaperone pifithrin-μ was mutation-specific. In contrast, lentiviral gene transfer of wild-type human SLC6A3 complementary DNA restored DAT activity and prevented neurodegeneration in all patient-derived mDA lines. To progress toward clinical translation, we used the knockout mouse model of DTDS that recapitulates human disease, exhibiting parkinsonism features, including tremor, bradykinesia, and premature death. Neonatal intracerebroventricular injection of human SLC6A3 using an adeno-associated virus (AAV) vector provided neuronal expression of human DAT, which ameliorated motor phenotype, life span, and neuronal survival in the substantia nigra and striatum, although off-target neurotoxic effects were seen at higher dosage. These were avoided with stereotactic delivery of AAV2.SLC6A3 gene therapy targeted to the midbrain of adult knockout mice, which rescued both motor phenotype and neurodegeneration, suggesting that targeted AAV gene therapy might be effective for patients with DTDS

The LRRK2-macroautophagy axis and its relevance to Parkinson's Disease

A wide variety of different functions and an impressive array of interactors have been associated with leucine-rich repeat kinase 2 (LRRK2) over the years. Here, I discuss the hypothesis that LRRK2 may be capable of interacting with different proteins at different times and places, therefore, controlling a plethora of diverse functions based on the different complexes formed. Among these, I will then focus on macroautophagy in the general context of the endolysosomal system. First, the relevance of autophagy in Parkinson's disease will be evaluated giving a brief overview of all the relevant Parkinson's disease genes; then, the association of LRRK2 with macroautophagy and the endolysosomal pathway will be analyzed based on the supporting literature

APOE and immunity: Research highlights

INTRODUCTION: At the Alzheimer's Association's APOE and Immunity virtual conference, held in October 2021, leading neuroscience experts shared recent research advances on and inspiring insights into the various roles that both the apolipoprotein E gene (APOE) and facets of immunity play in neurodegenerative diseases, including Alzheimer's disease and other dementias. METHODS: The meeting brought together more than 1200 registered attendees from 62 different countries, representing the realms of academia and industry. RESULTS: During the 4-day meeting, presenters illuminated aspects of the cross-talk between APOE and immunity, with a focus on the roles of microglia, triggering receptor expressed on myeloid cells 2 (TREM2), and components of inflammation (e.g., tumor necrosis factor α [TNFα]). DISCUSSION: This manuscript emphasizes the importance of diversity in current and future research and presents an integrated view of innate immune functions in Alzheimer's disease as well as related promising directions in drug development

Comparative protein interaction network analysis identifies shared and distinct functions for the human ROCO proteins

Signal transduction cascades governed by kinases and GTPases are a critical component of the command and control of cellular processes, with the precise outcome partly determined by direct protein‐protein interactions (PPIs). Here, we use the human ROCO proteins as a model for investigating PPI signalling events – taking advantage of the unique dual kinase/GTPase activities and scaffolding properties of these multidomain proteins. We report PPI networks that encompasses the human ROCO proteins, developed using two complementary approaches. First, using our recently developed weighted PPI network analysis (WPPINA) pipeline, a confidence‐weighted overview of validated ROCO protein interactors was obtained from peer‐reviewed literature. Second, novel ROCO PPIs were assessed experimentally via protein microarray screens. We compared the networks derived from these orthologous approaches to identify common elements within the ROCO protein interactome; functional enrichment analysis of this common core of the network identified stress response and cell projection organisation as shared functions within this protein family. Despite the presence of these commonalities, our results suggest that many unique interactors and therefore some specialised cellular roles have evolved for different members of the ROCO proteins. Overall, this multi‐approach strategy to increase the resolution of protein interaction networks represents a prototype for the utility of PPI data integration in understanding signalling biology

mTOR independent regulation of macroautophagy by Leucine Rich Repeat Kinase 2 via Beclin-1

Leucine rich repeat kinase 2 is a complex enzyme with both kinase and GTPase activities, closely linked to the pathogenesis of several human disorders including Parkinson’s disease, Crohn’s disease, leprosy and cancer. LRRK2 has been implicated in numerous cellular processes; however its physiological function remains unclear. Recent reports suggest that LRRK2 can act to regulate the cellular catabolic process of macroautophagy, although the precise mechanism whereby this occurs has not been identi ed. To investigate the signalling events through which LRRK2 acts to in uence macroautophagy, the mammalian target of rapamycin (mTOR)/Unc-51-like kinase 1 (ULK1) and Beclin-1/phosphatidylinositol 3-kinase (PI3K) pathways were evaluated in astrocytic cell models in the presence and absence of LRRK2 kinase inhibitors. Chemical inhibition of LRRK2 kinase activity resulted in the stimulation of macroautophagy in a non-canonical fashion, independent of mTOR and ULK1, but dependent upon the activation of Beclin 1-containing class III PI3-kinase

LRRK2 BAC transgenic rats develop progressive, L-DOPA-responsive motor impairment, and deficits in dopamine circuit function

Mutations in leucine-rich repeat kinase 2 (LRRK2) lead to late-onset, autosomal dominant Parkinson's disease, characterized by the degeneration of dopamine neurons of the substantia nigra pars compacta, a deficit in dopamine neurotransmission and the development of motor and non-motor symptoms. The most prevalent Parkinson's disease LRRK2 mutations are located in the kinase (G2019S) and GTPase (R1441C) encoding domains of LRRK2. To better understand the sequence of events that lead to progressive neurophysiological deficits in vulnerable neurons and circuits in Parkinson's disease, we have generated LRRK2 bacterial artificial chromosome transgenic rats expressing either G2019S or R1441C mutant, or wild-type LRRK2, from the complete human LRRK2 genomic locus, including endogenous promoter and regulatory regions. Aged (18–21 months) G2019S and R1441C mutant transgenic rats exhibit L-DOPA-responsive motor dysfunction, impaired striatal dopamine release as determined by fast-scan cyclic voltammetry, and cognitive deficits. In addition, in vivo recordings of identified substantia nigra pars compacta dopamine neurons in R1441C LRRK2 transgenic rats reveal an age-dependent reduction in burst firing, which likely results in further reductions to striatal dopamine release. These alterations to dopamine circuit function occur in the absence of neurodegeneration or abnormal protein accumulation within the substantia nigra pars compacta, suggesting that nigrostriatal dopamine dysfunction precedes detectable protein aggregation and cell death in the development of Parkinson's disease. In conclusion, our longitudinal deep-phenotyping provides novel insights into how the genetic burden arising from human mutant LRRK2 manifests as early pathophysiological changes to dopamine circuit function and highlights a potential model for testing Parkinson's therapeutics