4 research outputs found

    Single copy/knock-in models of ALS SOD1 in C. elegans suggest loss and gain of function have different contributions to cholinergic and glutamatergic neurodegeneration

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    Mutations in Cu/Zn superoxide dismutase 1 (SOD1) lead to Amyotrophic Lateral Sclerosis (ALS), a neurodegenerative disease that disproportionately affects glutamatergic and cholinergic motor neurons. Previous work with SOD1 overexpression models supports a role for SOD1 toxic gain of function in ALS pathogenesis. However, the impact of SOD1 loss of function in ALS cannot be directly examined in overexpression models. In addition, overexpression may obscure the contribution of SOD1 loss of function in the degeneration of different neuronal populations. Here, we report the first single-copy, ALS knock-in models in C. elegans generated by transposon- or CRISPR/Cas9- mediated genome editing of the endogenous sod-1 gene. Introduction of ALS patient amino acid changes A4V, H71Y, L84V, G85R or G93A into the C. elegans sod-1 gene yielded single-copy/knock-in ALS SOD1 models. These differ from previously reported overexpression models in multiple assays. In single-copy/knock-in models, we observed differential impact of sod-1 ALS alleles on glutamatergic and cholinergic neurodegeneration. A4V, H71Y, G85R, and G93A animals showed increased SOD1 protein accumulation and oxidative stress induced degeneration, consistent with a toxic gain of function in cholinergic motor neurons. By contrast, H71Y, L84V, and G85R lead to glutamatergic neuron degeneration due to sod-1 loss of function after oxidative stress. However, dopaminergic and serotonergic neuronal populations were spared in single-copy ALS models, suggesting a neuronal-subtype specificity previously not reported in invertebrate ALS SOD1 models. Combined, these results suggest that knock-in models may reproduce the neurotransmitter-type specificity of ALS and that both SOD1 loss and gain of toxic function differentially contribute to ALS pathogenesis in different neuronal populations.Peer reviewe

    The 11S proteasomal activator REGγ Impacts polyglutamine-expanded androgen receptor aggregation and toxicity in cell models of spinal and bulbar muscular atrophy

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    Spinal and bulbar muscular atrophy (SBMA) is a neurodegenerative disease that is caused by an expanded CAG repeat, which encodes a polyglutamine tract in the amino-terminal region of the androgen receptor (AR). Substantial data have illuminated the importance of the nucleus in SBMA pathogenesis; the expanded polyglutamine (polyQ) androgen receptor must reside in the nucleus, in the presence of ligand, in order to cause disease. Evidence that nuclear proteasomes inefficiently clear polyQ-expanded AR includes the finding that neuronal nuclear inclusions (NII), the pathological hallmark of SBMA, contain the amino-terminal portion of the AR, molecular chaperones, and components of the ubiquitin proteasome system (UPS). Here, we focus on how the nuclear 11S-proteasomal activator REGγ impacts polyQ-expanded AR metabolism. We reveal that REGã assumes either a proteasome binding-dependent or -independent role in the nucleus, depending on the cellular context. REGγ overexpression in a SBMA PC12 cell model increased polyQ-expanded AR aggregation and failed to protect cells from hormone-dependent toxicity through a proteasome binding-independent mechanism. Moreover, REGγ overexpression decreased polyQ-expanded AR polyubiquitylation and interaction with the E3 ligase MDM2. Preliminary genetic studies demonstrated that MDM2 overexpression may counter the REGγ aggregation effect, suggesting that REGγ acts as a competitor of the E3 ligase MDM2, influencing polyQ-expanded AR aggregation through its inhibition of AR polyubiquitylation and degradation. Conversely, we found that REGγ functions through its 11S-proteasomal activator role to rescue SBMA motor neurons from hormone-induced toxicity. How this direct proteasomal role works to protect SBMA motor neurons is currently unknown. Thus, our studies establish two biological roles of REGγ in impacting polyQ-expanded AR metabolism; exploration of the REGγ 11S-activator role may uncover novel targets for therapeutic advancement

    The 11S Proteasomal Activator REGγ Impacts Polyglutamine-Expanded Androgen Receptor Aggregation and Motor Neuron Viability through Distinct Mechanisms.

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    Spinal and bulbar muscular atrophy (SBMA) is caused by expression of a polyglutamine (polyQ)-expanded androgen receptor (AR). The inefficient nuclear proteasomal degradation of the mutant AR results in the formation of nuclear inclusions containing amino-terminal fragments of the mutant AR. PA28γ (also referred to as REGγ) is a nuclear 11S-proteasomal activator with limited proteasome activation capabilities compared to its cytoplasmic 11S (PA28α, PA28β) counterparts. To clarify the role of REGγ in polyQ-expanded AR metabolism, we carried out genetic and biochemical studies in cell models of SBMA. Overexpression of REGγ in a PC12 cell model of SBMA increased polyQ-expanded AR aggregation and contributed to polyQ-expanded AR toxicity in the presence of dihydrotestosterone (DHT). These effects of REGγ were independent of its association with the proteasome and may be due, in part, to the decreased binding of polyQ-expanded AR by the E3 ubiquitin-ligase MDM2. Unlike its effects in PC12 cells, REGγ overexpression rescued transgenic SBMA motor neurons from DHT-induced toxicity in a proteasome binding-dependent manner, suggesting that the degradation of a specific 11S proteasome substrate or substrates promotes motor neuron viability. One potential substrate that we found to play a role in mutant AR toxicity is the splicing factor SC35. These studies reveal that, depending on the cellular context, two biological roles for REGγ impact cell viability in the face of polyQ-expanded AR; a proteasome binding-independent mechanism directly promotes mutant AR aggregation while a proteasome binding-dependent mechanism promotes cell viability. The balance between these functions likely determines REGγ effects on polyQ-expanded AR-expressing cells
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