GPNMB confers risk for Parkinson's disease through interaction with α-synuclein.

Many risk loci for Parkinson's disease (PD) have been identified by genome-wide association studies (GWASs), but target genes and mechanisms remain largely unknown. We linked the GWAS-derived chromosome 7 locus (sentinel single-nucleotide polymorphism rs199347) to GPNMB through colocalization analyses of expression quantitative trait locus and PD risk signals, confirmed by allele-specific expression studies in the human brain. In cells, glycoprotein nonmetastatic melanoma protein B (GPNMB) coimmunoprecipitated and colocalized with α-synuclein (aSyn). In induced pluripotent stem cell-derived neurons, loss of GPNMB resulted in loss of ability to internalize aSyn fibrils and develop aSyn pathology. In 731 PD and 59 control biosamples, GPNMB was elevated in PD plasma, associating with disease severity. Thus, GPNMB represents a PD risk gene with potential for biomarker development and therapeutic targeting.

Increased expression of the frontotemporal dementia risk factor TMEM106B causes C9orf72-dependent alterations in lysosomes.

Frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) is an important cause of dementia in individuals under age 65. Common variants in the TMEM106B gene were previously discovered by genome-wide association to confer genetic risk for FTLD-TDP (p = 1 × 10-11, OR = 1.6). Furthermore, TMEM106B may act as a genetic modifier affecting age at onset and age at death in the Mendelian subgoup of FTLD-TDP due to expansions of the C9orf72 gene. Evidence suggests that TMEM106B variants increase risk for developing FTLD-TDP by increasing expression of Transmembrane Protein 106B (TMEM106B), a lysosomal protein. To further understand the functional role of TMEM106B in disease pathogenesis, we investigated the cell biological effects of increased TMEM106B expression. Here, we report that increased TMEM106B expression results in the appearance of a vacuolar phenotype in multiple cell types, including neurons. Concomitant with the development of this vacuolar phenotype, cells over-expressing TMEM106B exhibit impaired lysosomal acidification and degradative function, as well as increased cytotoxicity. We further identify a potential lysosomal sorting motif for TMEM106B and demonstrate that abrogation of sorting to lysosomes rescues TMEM106B-induced defects. Finally, we show that TMEM106B-induced defects are dependent on the presence of C9orf72, as knockdown of C9orf72 also rescues these defects. In sum, our results suggest that TMEM106B exerts its effects on FTLD-TDP disease risk through alterations in lysosomal pathways. Furthermore, TMEM106B and C9orf72 may interact in FTLD-TDP pathophysiology.

Pathologic and therapeutic implications for the cell biology of parkin.

Mutations in the E3 ligase parkin are the most common cause of autosomal recessive Parkinson's disease (PD), but it is believed that parkin dysfunction may also contribute to idiopathic PD. Since its discovery, parkin has been implicated in supporting multiple neuroprotective pathways, many revolving around the maintenance of mitochondrial health quality control and governance of cell survival. Recent advances across the structure, biochemistry, and cell biology of parkin have provided great insights into the etiology of parkin-linked and idiopathic PD and may ultimately generate novel therapeutic strategies to slow or halt disease progression. This review describes the various pathways in which parkin acts and the mechanisms by which parkin may be targeted for therapeutic intervention. This article is part of a Special Issue entitled 'Neuronal Protein'.

Adeno-associated virus serotype 8 (AAV8) delivery of recombinant A20 to skeletal muscle reduces pathological activation of nuclear factor (NF)-κB in muscle of mdx mice.

Duchenne muscular dystrophy (DMD) is a genetic muscle disease caused by the absence of a functional dystrophin protein. Lack of dystrophin protein disrupts the dystrophin-glycoprotein complex causing muscle membrane instability and degeneration. One of the secondary manifestations resulting from lack of functional dystrophin in muscle tissue is an increased level of cytokines that recruit inflammatory cells, leading to chronic upregulation of the nuclear factor (NF)-κB. Negative regulators of the classical NF-κB pathway improve muscle health in the mdx mouse model for DMD. We have previously shown in vitro that a negative regulator of the NF-κB pathway, A20, plays a role in muscle regeneration. Here, we show that overexpression of A20 by using a muscle-specific promoter delivered with an adeno-associated virus serotype 8 (AAV8) vector to the mdx mouse decreases activation of the NF-κB pathway in skeletal muscle. Recombinant A20 expression resulted in a reduction in number of fibers with centrally placed nuclei and a reduction in the number of T cells infiltrating muscle transduced with the AAV8-A20 vector. Taken together, we conclude that overexpression of A20 in mdx skeletal muscle provides improved muscle health by reduction of chronic inflammation and muscle degeneration. These results suggest A20 is a potential therapeutic target to ameliorate symptoms of DMD.

Deubiquitinating enzyme A20 negatively regulates NF-κB signaling in skeletal muscle in mdx mice.

Duchenne muscular dystrophy (DMD) is caused by the lack of a functional dystrophin protein that results in muscle fiber membrane disruption and, ultimately, degeneration. Regeneration of muscle fibers fails progressively, and muscle tissue is replaced with connective tissue. As a result, DMD causes progressive limb muscle weakness and cardiac and respiratory failure. The absence of dystrophin from muscle fibers triggers the chronic activation of the nuclear factor of kappa B (NF-κB). Chronic activation of NF-κB in muscle leads to infiltration of macrophages, up-regulation of the ubiquitin-proteosome system, and down-regulation of the helix-loop-helix muscle regulatory factor, MyoD. These processes, triggered by NF-κB activation, promote muscle degeneration and failure of muscle regeneration. A20 (TNFAIP3) is a critical negative regulator of NF-κB. In this study, we characterize the role of A20 in regulating NF-κB activation in skeletal muscle, identifying a novel role in muscle regeneration. A20 is highly expressed in regenerating muscle fibers, and knockdown of A20 impairs muscle differentiation in vitro, which suggests that A20 expression is critically important for regeneration of dystrophic muscle tissue. Furthermore, down-regulation of the classic pathway of NF-κB activation is associated with up-regulation of the alternate pathway in regenerating muscle fibers, suggesting a mechanism by which A20 promotes muscle regeneration. These results demonstrate the important role of A20 in muscle fiber repair and suggest the potential of A20 as a therapeutic target to ameliorate the pathology and clinical symptoms of DMD.

A potential role of distinctively delayed blood clearance of recombinant adeno-associated virus serotype 9 in robust cardiac transduction.

Recombinant adeno-associated virus serotype 9 (rAAV9) vectors show robust in vivo transduction by a systemic approach. It has been proposed that rAAV9 has enhanced ability to cross the vascular endothelial barriers. However, the scientific basis of systemic administration of rAAV9 and its transduction mechanisms have not been fully established. Here, we show indirect evidence suggesting that capillary walls still remain as a significant barrier to rAAV9 in cardiac transduction but not so in hepatic transduction in mice, and the distinctively delayed blood clearance of rAAV9 plays an important role in overcoming this barrier, contributing to robust cardiac transduction. We find that transvascular transport of rAAV9 in the heart is a capacity-limited slow process and occurs in the absence of caveolin-1, the major component of caveolae that mediate endothelial transcytosis. In addition, a reverse genetic study identifies the outer region of the icosahedral threefold capsid protrusions as a potential culprit for rAAV9's delayed blood clearance. These results support a model in which the delayed blood clearance of rAAV9 sustains the capacity-limited slow transvascular vector transport and plays a role in mediating robust cardiac transduction, and provide important implications in AAV capsid engineering to create new rAAV variants with more desirable properties.

Kinesin-2 differentially regulates the anterograde axonal transports of acetylcholinesterase and choline acetyltransferase in Drosophila.

Choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) are involved in acetylcholine synthesis and degradation at pre- and postsynaptic compartments, respectively. Here we show that their anterograde transport in Drosophila larval ganglion is microtubule-dependent and occurs in two different time profiles. AChE transport is constitutive while that of ChAT occurs in a brief pulse during third instar larva stage. Mutations in the kinesin-2 motor subunit Klp64D and separate siRNA-mediated knock-outs of all the three kinesin-2 subunits disrupt the ChAT and AChE transports, and these antigens accumulate in discrete nonoverlapping punctae in neuronal cell bodies and axons. Quantification analysis further showed that mutations in Klp64D could independently affect the anterograde transport of AChE even before that of ChAT. Finally, ChAT and AChE were coimmunoprecipitated with the kinesin-2 subunits but not with each other. Altogether, these suggest that kinesin-2 independently transports AChE and ChAT within the same axon. It also implies that cargo availability could regulate the rate and frequency of transports by kinesin motors.