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1.  Regulation of endosomal motility and degradation by amyotrophic lateral sclerosis 2/alsin 
Molecular Brain  2009;2:23.
Dysfunction of alsin, particularly its putative Rab5 guanine-nucleotide-exchange factor activity, has been linked to one form of juvenile onset recessive familial amyotrophic lateral sclerosis (ALS2). Multiple lines of alsin knockout (ALS2-/-) mice have been generated to model this disease. However, it remains elusive whether the Rab5-dependent endocytosis is altered in ALS2-/- neurons. To directly examine the Rab5-mediated endosomal trafficking in ALS2-/- neurons, we introduced green fluorescent protein (GFP)-tagged Rab5 into cultured hippocampal neurons to monitor the morphology and motility of Rab5-associated early endosomes. Here we report that Rab5-mediated endocytosis was severely altered in ALS2-/-neurons. Excessive accumulation of Rab5-positive vesicles was observed in ALS2-/- neurons, which correlated with a significant reduction in endosomal motility and augmentation in endosomal conversion to lysosomes. Consequently, a significant increase in endosome/lysosome-dependent degradation of internalized glutamate receptors was observed in ALS2-/- neurons. These phenotypes closely resembled the endosomal trafficking abnormalities induced by a constitutively active form of Rab5 in wild-type neurons. Therefore, our findings reveal a negatively regulatory mechanism of alsin in Rab5-mediated endosomal trafficking, suggesting that enhanced endosomal degradation in ALS2-/- neurons may underlie the pathogenesis of motor neuron degeneration in ALS2 and related motor neuron diseases.
doi:10.1186/1756-6606-2-23
PMCID: PMC2724476  PMID: 19630956
2.  Progressive Behavioral Deficits in DJ-1 Deficient Mice are Associated with Normal Nigrostriatal Function 
Neurobiology of disease  2007;29(3):505-514.
Loss-of-function mutations in the DJ-1 gene account for an autosomal recessive form of Parkinson’s disease (PD). To investigate the physiological functions of DJ-1 in vivo, we generated DJ-1 knockout (DJ-1-/-) mice. Younger (< 1year) DJ-1 -/- mice were hypoactive and had mild gait abnormalities. Older DJ-1-/-, however, showed decreased bodyweight and grip strength, and more severe gait irregularities compared to wild-type littermates. The basal level of extracellular dopamine, evoked dopamine release and dopamine receptor D2 sensitivity appeared normal in the striatum of DJ-1-/- mice, which was consistent with similar results between DJ-1-/- and controls in behavioral paradigms specific for the dopaminergic system. An examination of spinal cord, nerve and muscle tissues failed to identify any pathological changes that were consistent with the noted motor deficits. Taken together, our findings suggest that loss of DJ-1 leads to progressive behavioral changes without significant alterations in nigrostriatal dopaminergic and spinal motor systems.
doi:10.1016/j.nbd.2007.11.011
PMCID: PMC2271119  PMID: 18187333
DJ-1; knockout mouse; Parkinson’s disease; dopamine; striatum; spinal cord; muscle; motor behavior
3.  The Chaperone Activity of Heat Shock Protein 90 Is Critical for Maintaining the Stability of Leucine-Rich Repeat Kinase 2 
Parkinson’s disease (PD), a progressive neurodegenerative disease characterized by bradykinesia, rigidity, and resting tremor, is the most common neurodegenerative movement disorder. Although the majority of PD cases are sporadic, some are inherited, including those caused by leucine-rich repeat kinase 2 (LRRK2) mutations. The substitution of serine for glycine at position 2019 (G2019S) in the kinase domain of LRRK2 represents the most prevalent genetic mutation in both familial and apparently sporadic cases of PD. Because mutations in LRRK2 are likely associated with a toxic gain of function, destabilization of LRRK2 may be a novel way to limit its detrimental effects. Here we show that LRRK2 forms a complex with heat shock protein 90 (Hsp90) in vivo and that inhibition of Hsp90 disrupts the association of Hsp90 with LRRK2 and leads to proteasomal degradation of LRRK2. Hsp90 inhibitors may therefore limit the mutant LRRK2-elicited toxicity to neurons. As a proof of principle, we show that Hsp90 inhibitors rescue the axon growth retardation caused by overexpression of the LRRK2 G2019S mutation in neurons. Therefore, inhibition of LRRK2 kinase activity can be achieved by blocking Hsp90-mediated chaperone activity and Hsp90 inhibitors may serve as potential anti-PD drugs.
doi:10.1523/JNEUROSCI.0185-08.2008
PMCID: PMC2564280  PMID: 18367605
Hsp90; LRRK2; G2019S; Parkinson’s disease; protein degradation; chaperone
4.  ALS2/Alsin Knockout Mice and Motor Neuron Diseases 
Neuro-degenerative diseases  2008;5(6):359-366.
Autosomal recessive mutations in the ALS2 gene have been linked to juvenile-onset amyotrophic lateral sclerosis (ALS2), primary lateral sclerosis and juvenile-onset ascending hereditary spastic paraplegia. Except for two recently identified missense mutations, all other mutations in the ALS2 gene lead to a premature stop codon and likely abrogate all the potential functions of alsin, the protein encoded by the ALS2 gene. To study the pathologic mechanisms of ALS2 deficiency, four different lines of ALS2 knockout (ALS2−/−) mice have been generated by independent groups. The loss of ALS2/alsin does not have a drastic effect on the survival or function of motor neurons in mice. However, subtle deficits observed in the behavior and pathology of these mice have aided in our understanding of the relationship between alsin and motor neuron dysfunction. In this review, we summarize and reconcile major findings of ALS2−/− mice and attempt to place these results within the larger context of modeling recessive movement disorders in mice.
doi:10.1159/000151295
PMCID: PMC2556598  PMID: 18714162
Amyotrophic lateral sclerosis; ALS2; Alsin; Knockout mice; Mouse model; Guanine nucleotide exchange factor; Primary lateral sclerosis; Hereditary spastic paraplegia
5.  Amyotrophic Lateral Sclerosis 2-Deficiency Leads to Neuronal Degeneration in Amyotrophic Lateral Sclerosis through Altered AMPA Receptor Trafficking 
Amyotrophic lateral sclerosis (ALS), the most common adult-onset motor neuron disease is caused by a selective loss of motor neurons. One form of juvenile onset autosomal recessive ALS (ALS2) has been linked to the loss of function of the ALS2 gene. The pathogenic mechanism of ALS2-deficiency, however, remains unclear. To further understand the function of alsin that is encoded by the full-length ALS2 gene, we screened proteins interacting with alsin. Here, we report that alsin interacted with glutamate receptor interacting protein 1 (GRIP1) both in vitro and in vivo, and colocalized with GRIP1 in neurons. In support of the physiological interaction between alsin and GRIP1, the subcellular distribution of GRIP1 was altered in ALS2-/- spinal motor neurons, which correlates with a significant reduction of AMPA-type glutamate receptor subunit 2 (GluR2) at the synaptic/cell surface of ALS2-/- neurons. The decrease of calcium-impermeable GluR2-containing AMPA receptors at the cell/synaptic surface rendered ALS2-/- neurons more susceptible to glutamate receptor-mediated neurotoxicity. Our findings reveal a novel function of alsin in AMPA receptor trafficking and provide a novel pathogenic link between ALS2-deficiency and motor neuron degeneration, suggesting a protective role of alsin in maintaining the survival of motor neurons.
doi:10.1523/JNEUROSCI.2084-06.2006
PMCID: PMC2556290  PMID: 17093100
ALS2; knock-out mouse; motor neuron; GRIP1; AMPA receptor; excitotoxicity
6.  ALS2/Alsin Knockout Mice and Motor Neuron Diseases 
Neuro-Degenerative Diseases  2008;5(6):359-366.
Autosomal recessive mutations in the ALS2 gene have been linked to juvenile-onset amyotrophic lateral sclerosis (ALS2), primary lateral sclerosis and juvenile-onset ascending hereditary spastic paraplegia. Except for two recently identified missense mutations, all other mutations in the ALS2 gene lead to a premature stop codon and likely abrogate all the potential functions of alsin, the protein encoded by the ALS2 gene. To study the pathologic mechanisms of ALS2 deficiency, four different lines of ALS2 knockout (ALS2–/–) mice have been generated by independent groups. The loss of ALS2/alsin does not have a drastic effect on the survival or function of motor neurons in mice. However, subtle deficits observed in the behavior and pathology of these mice have aided in our understanding of the relationship between alsin and motor neuron dysfunction. In this review, we summarize and reconcile major findings of ALS2–/– mice and attempt to place these results within the larger context of modeling recessive movement disorders in mice.
doi:10.1159/000151295
PMCID: PMC2556598  PMID: 18714162
Amyotrophic lateral sclerosis; ALS2; Alsin; Knockout mice; Mouse model; Guanine nucleotide exchange factor; Primary lateral sclerosis; Hereditary spastic paraplegia
7.  The G59S Mutation in p150glued Causes Dysfunction of Dynactin in Mice 
The G59S missense mutation at the conserved microtubule-binding domain of p150glued, a major component of dynein/dynactin complex, has been linked to an autosomal dominant form of motor neuron disease (MND). To study how this mutation affects the function of the dynein/dynactin complex and contributes to motor neuron degeneration, we generated p150glued G59S knock-in mice. We found that the G59S mutation destabilizes p150glued and disrupts the function of dynein/dynactin complex, resulting in early embryonic lethality of homozygous knock-in mice. Heterozygous knock-in mice, which developed normally, displayed MND-like phenotypes after 10 months of age, including excessive accumulation of cytoskeletal and synaptic vesicle proteins at neuromuscular junctions, loss of spinal motor neurons, increase of reactive astrogliosis, and shortening of gait compared with wild-type littermates and age-matched p150glued heterozygous knock-out mice. Our findings indicate that the G59S mutation in p150glued abrogates the normal function of p150glued and accelerates motor neuron degeneration.
doi:10.1523/JNEUROSCI.4226-07.2007
PMCID: PMC2367233  PMID: 18094236
dynactin; dynein; p150glued; motor neuron disease; mouse model; ALS
8.  Alsin and the Molecular Pathways of Amyotrophic Lateral Sclerosis 
Molecular neurobiology  2007;36(3):224-231.
Autosomal recessive mutations in the ALS2 gene lead to a clinical spectrum of motor dysfunction including juvenile onset amyotrophic lateral sclerosis (ALS2), primary lateral sclerosis, and hereditary spastic paraplegia. The 184-kDa alsin protein, encoded by the full-length ALS2 gene, contains three different guanine-nucleotide-exchange factor-like domains, which may play a role in the etiology of the disease. Multiple in vitro biochemical and cell biology assays suggest that alsin dysfunction affects endosome trafficking through a Rab5 small GTPase family-mediated mechanism. Four ALS2-deficient mouse models have been generated by different groups and used to study the behavioral and pathological impact of alsin deficiency. These mouse models largely fail to recapitulate hallmarks of motor neuron disease, but the subtle deficits that are observed in behavior and pathology have aided in our understanding of the relationship between alsin and motor dysfunction. In this review, we summarize recent clinical and molecular reports regarding alsin and attempt to place these results within the larger context of motor neuron disease.
doi:10.1007/s12035-007-0034-x
PMCID: PMC2364715  PMID: 17955197
Amyotrophic lateral sclerosis (ALS); ALS2; Alsin; Rab5; Mouse model; Guanine-nucleotide-exchange factor; Primary lateral sclerosis; Hereditary spastic paraplegia
9.  Deletion at ITPR1 Underlies Ataxia in Mice and Spinocerebellar Ataxia 15 in Humans 
PLoS Genetics  2007;3(6):e108.
We observed a severe autosomal recessive movement disorder in mice used within our laboratory. We pursued a series of experiments to define the genetic lesion underlying this disorder and to identify a cognate disease in humans with mutation at the same locus. Through linkage and sequence analysis we show here that this disorder is caused by a homozygous in-frame 18-bp deletion in Itpr1 (Itpr1Δ18/Δ18), encoding inositol 1,4,5-triphosphate receptor 1. A previously reported spontaneous Itpr1 mutation in mice causes a phenotype identical to that observed here. In both models in-frame deletion within Itpr1 leads to a decrease in the normally high level of Itpr1 expression in cerebellar Purkinje cells. Spinocerebellar ataxia 15 (SCA15), a human autosomal dominant disorder, maps to the genomic region containing ITPR1; however, to date no causal mutations had been identified. Because ataxia is a prominent feature in Itpr1 mutant mice, we performed a series of experiments to test the hypothesis that mutation at ITPR1 may be the cause of SCA15. We show here that heterozygous deletion of the 5′ part of the ITPR1 gene, encompassing exons 1–10, 1–40, and 1–44 in three studied families, underlies SCA15 in humans.
Author Summary
We have identified a spontaneous in-frame deletion mutation in the gene Itpr1 that causes a recessive movement disorder in mice. In an attempt to define whether any similar disease occurs in humans we performed a literature search for diseases linked to the human chromosomal region containing ITPR1. We identified the disease spinocerebellar ataxia 15 as linked to this region. High-density genomic analysis of affected members from three families revealed that disease in these patients was caused by deletion of a large portion of the region containing ITPR1. We show here that this mutation results in a dramatic reduction in ITPR1 in cells from these patients. These data show convincingly that ITPR1 deletion underlies spinocerebellar ataxia 15 in humans.
doi:10.1371/journal.pgen.0030108
PMCID: PMC1892049  PMID: 17590087
10.  Deletion at ITPR1 Underlies Ataxia in Mice and Spinocerebellar Ataxia 15 in Humans 
PLoS Genetics  2007;3(6):e108.
We observed a severe autosomal recessive movement disorder in mice used within our laboratory. We pursued a series of experiments to define the genetic lesion underlying this disorder and to identify a cognate disease in humans with mutation at the same locus. Through linkage and sequence analysis we show here that this disorder is caused by a homozygous in-frame 18-bp deletion in Itpr1 (Itpr1Δ18/Δ18), encoding inositol 1,4,5-triphosphate receptor 1. A previously reported spontaneous Itpr1 mutation in mice causes a phenotype identical to that observed here. In both models in-frame deletion within Itpr1 leads to a decrease in the normally high level of Itpr1 expression in cerebellar Purkinje cells. Spinocerebellar ataxia 15 (SCA15), a human autosomal dominant disorder, maps to the genomic region containing ITPR1; however, to date no causal mutations had been identified. Because ataxia is a prominent feature in Itpr1 mutant mice, we performed a series of experiments to test the hypothesis that mutation at ITPR1 may be the cause of SCA15. We show here that heterozygous deletion of the 5′ part of the ITPR1 gene, encompassing exons 1–10, 1–40, and 1–44 in three studied families, underlies SCA15 in humans.
Author Summary
We have identified a spontaneous in-frame deletion mutation in the gene Itpr1 that causes a recessive movement disorder in mice. In an attempt to define whether any similar disease occurs in humans we performed a literature search for diseases linked to the human chromosomal region containing ITPR1. We identified the disease spinocerebellar ataxia 15 as linked to this region. High-density genomic analysis of affected members from three families revealed that disease in these patients was caused by deletion of a large portion of the region containing ITPR1. We show here that this mutation results in a dramatic reduction in ITPR1 in cells from these patients. These data show convincingly that ITPR1 deletion underlies spinocerebellar ataxia 15 in humans.
doi:10.1371/journal.pgen.0030108
PMCID: PMC1892049  PMID: 17590087

Results 1-10 (10)