356 research outputs found
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
Sequential extraction of soluble and insoluble alpha-synuclein from Parkinsonian brains
Alpha-synuclein (α-syn) protein is abundantly expressed mainly within neurons, and exists in a number of different forms - monomers, tetramers, oligomers and fibrils. During disease, α-syn undergoes conformational changes to form oligomers and high molecular weight aggregates that tend to make the protein more insoluble. Abnormally aggregated α-syn is a neuropathological feature of Parkinsonâs disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). Biochemical characterisation and analysis of insoluble α-syn using buffers with increasing detergent strength and high-speed ultracentrifugation provides a powerful tool to determine the development of α-syn pathology associated with disease progression. This protocol describes the isolation of increasingly insoluble/aggregated αâsyn from post-mortem human brain tissue. This methodology can be adapted with modifications to studies of normal and abnormal α-syn biology in transgenic animal models harbouring different α-syn mutations as well as in other neurodegenerative diseases that feature aberrant fibrillar deposits of proteins related to their respective pathologies
A conceptual model for identifying the risk susceptibility of urban green spaces using geo-spatial techniques
The presence of heterogeneous nuclear ribonucleoproteins in frontotemporal lobar degeneration with FUS positive inclusions
Frontotemporal lobar degeneration with fused in sarcomaâpositive inclusions (FTLD-FUS) is a disease with unknown cause. Transportin 1 is abundantly found in FUS-positive inclusions and responsible for the nuclear import of the FET proteins of which FUS is a member. The presence of all FET proteins in pathological inclusions suggests a disturbance of transportin 1âmediated nuclear import. FUS also belongs to the heterogeneous nuclear ribonucleoprotein (hnRNP) protein family. We investigated whether hnRNP proteins are associated with FUS pathology implicating dysfunctional nuclear export in the pathogenesis of FTLD-FUS. hnRNP proteins were investigated in affected brain regions in FTLD-FUS using immunohistochemistry, biochemical analysis, and the expression analysis. We demonstrated the presence of several hnRNP proteins in pathological inclusions including neuronal cytoplasmic inclusions and dystrophic neurites. The biochemical analysis revealed a shift in the location of hnRNP A1 from the nucleus to the cytoplasm. The expression analysis revealed an increase in several hnRNP proteins in FTLD-FUS. These results implicate a wider dysregulation of movement between intracellular compartments, than mechanisms only affecting the nuclear import of FUS proteins
An operator-induced conformational change in the C-terminal domain of the λ repressor
4,4'-bis(1-anilino-8-naphthalenesulfonic acid (Bis-ANS), an environment-sensitive fluorescent probe for hydrophobic region of proteins, binds specifically to the C-terminal domain of λ repressor. The binding is characterized by positive cooperativity, the magnitude of which is dependent on protein concentration in the concentration range where dimeric repressor aggregates to a tetramer. In this range, positive cooperativity becomes more pronounced at higher protein concentrations. This suggests a preferential binding of Bis-ANS to the dimeric form of the repressor. Binding of single operator OR1 to the N-terminal domain of the repressor causes enhancement of fluorescence of the C-terminal domain bound Bis-ANS. The binding of single operator OR1 also leads to quenching of fluorescence of tryptophan residues, all of which are located in the hinge or the C-terminal domain. Thus two different fluorescent probes indicate an operator-induced conformational change which affects the C-terminal domain. The significance of this conformational change with respect to the function of λ repressor has been discussed
In silico comparative analysis of LRRK2 interactomes from brain, kidney and lung
Mutations in LRRK2 are the most frequent cause of familial Parkinsonâs disease (PD), with common LRRK2 non-coding variants also acting as risk factors for idiopathic PD. Currently, therapeutic agents targeting LRRK2 are undergoing advanced clinical trials in humans, however, it is important to understand the wider implications of LRRK2 targeted treatments given that LRRK2 is expressed in diverse tissues including the brain, kidney and lungs. This presents challenges to treatment in terms of effects on peripheral organ functioning, thus, protein interactors of LRRK2 could be targeted in lieu to optimize therapeutic effects. Herein an in-silico analysis of LRRK2 direct interactors in brain tissue from various brain regions was conducted along with a comparative analysis of the LRRK2 interactome in the brain, kidney, and lung tissues. This was carried out based on curated proteinâprotein interaction (PPI) data from protein interaction databases such as HIPPIE, human gene/protein expression databases and Gene ontology (GO) enrichment analysis using Bingo. Seven targets (MAP2K6, MATK, MAPT, PAK6, SH3GL2, CDC42EP3 and CHGB) were found to be viable objectives for LRRK2 based investigations for PD that would have minimal impact on optimal functioning within peripheral organs. Specifically, MAPT, CHGB, PAK6, and SH3GL2 interacted with LRRK2 in the brain and kidney but not in lung tissue whilst LRRK2-MAP2K6 interacted only in the cerebellum and MATK-LRRK2 interaction was absent in kidney tissues. CDC42EP3 expression levels were low in brain tissues compared to kidney/lung. The results of this computational analysis suggest new avenues for experimental investigations towards LRRK2-targeted therapeutics
Pathological relevance of post-translationally modified alpha-synuclein (pSer87, pSer129, nTyr39) in idiopathic Parkinsonâs disease and Multiple System Atrophy
Aggregated alpha-synuclein (a-synuclein) is the main component of Lewy bodies (LBs), Lewy neurites (LNs), and glial cytoplasmic inclusions (GCIs), which are pathological hallmarks of idiopathic Parkinsonâs disease (IPD) and multiple system atrophy (MSA), respectively. Initiating factors that culminate in forming LBs/LNs/GCIs remain elusive. Several species of a-synuclein exist, including phosphorylated and nitrated forms. It is unclear which a-synuclein post-translational modifications (PTMs) appear within aggregates throughout disease pathology. Herein we aimed to establish the predominant a-synuclein PTMs in post-mortem IPD and MSA pathology using immunohistochemistry. We examined the patterns of three a-synuclein PTMs (pS87, pS129, nY39) simultaneously in pathology- affected regions of 15 PD, 5 MSA, 6 neurologically normal controls. All antibodies recognized LBs, LNs, and GCIs, albeit to a variable extent. pS129 a-synuclein antibody was particularly immunopositive for LNs and synaptic dot-like structures followed by nY39 a- synuclein antibody. GCIs, neuronal inclusions, and small threads were positive for nY39 a- synuclein in MSA. Quantification of the LB scores revealed that pS129 a-synuclein was the dominant and earliest a-synuclein PTM followed by nY39 a-synuclein, while lower amounts of pSer87 a-synuclein appeared later in disease progression in PD. These results may have implications for novel biomarker and therapeutic developments
Trafficking of the glutamate transporter is impaired in LRRK2 related Parkinson's disease
The Excitatory Amino Acid Transporter 2 (EAAT2) accounts for 80 % of brain glutamate clearance and is mainly expressed in astrocytic perisynaptic processes. EAAT2 function is finely regulated by endocytic events, recycling to the plasma membrane and degradation. Noteworthy, deficits in EAAT2 have been associated with neuronal excitotoxicity and neurodegeneration. In this study, we show that EAAT2 trafficking is impaired by the leucine-rich repeat kinase 2 (LRRK2) pathogenic variant G2019S, a common cause of late-onset familial Parkinsonâs disease (PD). In LRRK2 G2019S human brains and experimental animal models, EAAT2 protein levels are significantly decreased, which is associated with elevated gliosis. The decreased expression of the transporter correlates with its reduced functionality in mouse LRRK2 G2019S purified astrocytic terminals and in Xenopus laevis oocytes expressing human LRRK2 G2019S. In Lrrk2 G2019S knockin mouse brain, the correct surface localization of the endogenous transporter is impaired, resulting in its interaction with a plethora of endo-vesicular proteins. Mechanistically, we report that pathogenic LRRK2 kinase activity delays the recycling of the transporter to the plasma membrane, causing its intracellular relocalization and degradation. Taken together, our results demonstrate that pathogenic LRRK2 interferes with the physiology of EAAT2, pointing to extracellular glutamate overload as a possible contributor to neurodegeneration in PD
Survival of orbiting in Ne (7 - 10 MeV/nucleon) + C reactions
The inclusive energy distributions of fragments with Z 3 emitted from
the bombardment of C by Ne beams with incident energies between
145 and 200 MeV have been measured in the angular range
10 - 50. Damped fragment yields in all cases have been found to
be characteristic of emission from fully energy equilibrated composites; for B,
C fragments, average Q-values, , were independent of the centre of mass
emission angle (), and the angular distributions followed
1/sin like variation, signifying long life times of the
emitting di-nuclear systems. Total yields of these fragments have been found to
be much larger compared to the standard statistical model predictions of the
same. This may be indicative of the survival of orbiting like process in
C + Ne system at these energies.Comment: 7 pages, 5 figures, accepted for publication in Phys. Rev. C (Rapid
Communication
Characterization of fragment emission in ^{20}Ne (7 - 10 MeV/nucleon) + ^{12}C reactions
The inclusive energy distributions of the complex fragments (3 Z
7) emitted from the bombardment of ^{12}C by ^{20}Ne beams with incident
energies between 145 and 200 MeV have been measured in the angular range
10 50^{o}. Damped fragment yields in all the cases
have been found to be the characteristic of emission from fully energy
equilibrated composites. The binary fragment yields are compared with the
standard statistical model predictions. Enhanced yields of entrance channel
fragments (5 Z 7) indicate the survival of orbiting-like process
in ^{20}Ne + ^{12}C system at these energies.Comment: 18 pages, 13 figure
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