Modeling Protein Aggregation and the Heat Shock Response in ALS iPSC-Derived Motor Neurons

Abstract Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are now widely considered to be part of a disease spectrum with the identification of common pathological features and genetic causes. However, despite these advances, there remains no effective therapy for these conditions. In this study we demonstrate that mice expressing mutant valosin containing protein (VCP) develop an ALS/FTD-like phenotype in the spinal cord and brain, and treatment with arimoclomol, a pharmacological amplifier of the cytoprotective heat shock response ameliorates this phenotype. Moreover, the beneficial effects of arimoclomol are seen in both fibroblasts and iPSC-derived motor neurons from patients. Importantly, we show the pathological changes targeted by arimoclomol in our experimental models are present in post-mortem FTD patient tissue. Together with previous data demonstrating the efficacy of arimoclomol in SOD1-ALS models, our findings suggest that arimoclomol may have therapeutic potential not only in non-SOD1 ALS but also for the treatment of FTD.

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Depending on the stress, histone deacetylase inhibitors act as heat shock protein co-inducers in motor neurons and potentiate arimoclomol, exerting neuroprotection through multiple mechanisms in ALS models

Cell Stress and Chaperones ◽

10.1007/s12192-019-01064-1 ◽

2020 ◽

Vol 25 (1) ◽

pp. 173-191 ◽

Cited By ~ 7

Author(s):

Rachel Kuta ◽

Nancy Larochelle ◽

Mario Fernandez ◽

Arun Pal ◽

Sandra Minotti ◽

...

Keyword(s):

Heat Shock ◽

Histone Deacetylase ◽

Motor Neurons ◽

Heat Shock Response ◽

Hdac Inhibitors ◽

Dependent Manner ◽

Hdac Inhibition ◽

Proteotoxic Stress ◽

Shock Response ◽

Stress Dependent

AbstractUpregulation of heat shock proteins (HSPs) is an approach to treatment of neurodegenerative disorders with impaired proteostasis. Many neurons, including motor neurons affected in amyotrophic lateral sclerosis (ALS), are relatively resistant to stress-induced upregulation of HSPs. This study demonstrated that histone deacetylase (HDAC) inhibitors enable the heat shock response in cultured spinal motor neurons, in a stress-dependent manner, and can improve the efficacy of HSP-inducing drugs in murine spinal cord cultures subjected to thermal or proteotoxic stress. The effect of particular HDAC inhibitors differed with the stress paradigm. The HDAC6 (class IIb) inhibitor, tubastatin A, acted as a co-inducer of Hsp70 (HSPA1A) expression with heat shock, but not with proteotoxic stress induced by expression of mutant SOD1 linked to familial ALS. Certain HDAC class I inhibitors (the pan inhibitor, SAHA, or the HDAC1/3 inhibitor, RGFP109) were HSP co-inducers comparable to the hydroxyamine arimoclomol in response to proteotoxic stress, but not thermal stress. Regardless, stress-induced Hsp70 expression could be enhanced by combining an HDAC inhibitor with either arimoclomol or with an HSP90 inhibitor that constitutively induced HSPs. HDAC inhibition failed to induce Hsp70 in motor neurons expressing ALS-linked mutant FUS, in which the heat shock response was suppressed; yet SAHA, RGFP109, and arimoclomol did reduce loss of nuclear FUS, a disease hallmark, and HDAC inhibition rescued the DNA repair response in iPSC-derived motor neurons carrying the FUSP525Lmutation, pointing to multiple mechanisms of neuroprotection by both HDAC inhibiting drugs and arimoclomol.

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Neurodegenerative disease-associated protein aggregates are poor inducers of the heat shock response in neuronal-like cells

10.1101/2020.01.06.896654 ◽

2020 ◽

Cited By ~ 1

Author(s):

R. San Gil ◽

D. Cox ◽

L. McAlary ◽

T. Berg ◽

A. K. Walker ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Neurodegenerative Diseases ◽

Protein Aggregation ◽

Neurodegenerative Disease ◽

Heat Shock Response ◽

Flow Cytometric Analysis ◽

Protein Aggregates ◽

Shock Response ◽

Shock Proteins

AbstractProtein aggregation that results in the formation of inclusions is strongly correlated with neuronal death and is a pathological hallmark common to many neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and Huntington’s disease. Cells are thought to dramatically up-regulate the levels of heat shock proteins during periods of cellular stress via induction of the heat shock response (HSR). Heat shock proteins are well-characterised molecular chaperones that interact with aggregation-prone proteins to either stabilise, refold, or traffic protein for degradation. The reason why heat shock proteins are unable to maintain the solubility of particular proteins in neurodegenerative disease is unknown. We sought to determine whether neurodegenerative disease-associated protein aggregates can induce the HSR. Here, we generated a neuroblastoma cell line that expresses a fluorescent reporter under conditions of HSR induction, for example heat shock. Using these cells, we show that the HSR is not induced by exogenous treatment with aggregated forms of Parkinson’s disease-associated α-synuclein or the ALS-associated G93A mutant of superoxide dismutase-1 (SOD1G93A). Furthermore, flow cytometric analysis revealed that intracellular expression of SOD1G93A or a pathogenic form of polyQ-expanded huntingtin (Htt72Q), similarly, results in no or low induction of the HSR. In contrast, expression of a non-pathogenic but aggregation-prone form of firefly luciferase (Fluc) did induce an HSR in a significantly greater proportion of cells. Finally, we show that HSR induction is dependent on the intracellular levels of the aggregation-prone proteins, but the pathogenic proteins (SOD1G93A and Htt72Q) elicit a significantly lower HSR compared to the non-pathogenic proteins (Fluc). These results suggest that pathogenic proteins either evade detection or impair induction of the HSR in neuronal-like cells. Therefore, defective HSR induction may facilitate the initiation of protein aggregation leading to inclusion formation in neurodegenerative diseases.

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Heat Shock Response: Cellular and Molecular Responses to Stress, Misfolded Proteins, and Diseases Associated with Protein Aggregation

Wiley Encyclopedia of Molecular Medicine ◽

10.1002/0471203076.emm1261 ◽

2002 ◽

Author(s):

Ellen A. A. Nollen ◽

Richard I. Morimoto

Keyword(s):

Heat Shock ◽

Protein Aggregation ◽

Heat Shock Response ◽

Misfolded Proteins ◽

Molecular Responses ◽

Shock Response

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Histamine Is an Inducer of the Heat Shock Response in SOD1-G93A Models of ALS

International Journal of Molecular Sciences ◽

10.3390/ijms20153793 ◽

2019 ◽

Vol 20 (15) ◽

pp. 3793 ◽

Cited By ~ 2

Author(s):

Savina Apolloni ◽

Francesca Caputi ◽

Annabella Pignataro ◽

Susanna Amadio ◽

Paola Fabbrizio ◽

...

Keyword(s):

Heat Shock ◽

Motor Neuron ◽

Motor Neurons ◽

Heat Shock Response ◽

Type I ◽

Spine Density ◽

Shock Response ◽

Anti Inflammatory

(1) Background: Amyotrophic lateral sclerosis (ALS) is a multifactorial non-cell autonomous disease where activation of microglia and astrocytes largely contributes to motor neurons death. Heat shock proteins have been demonstrated to promote neuronal survival and exert a strong anti-inflammatory action in glia. Having previously shown that the pharmacological increase of the histamine content in the central nervous system (CNS) of SOD1-G93A mice decreases neuroinflammation, reduces motor neuron death, and increases mice life span, here we examined whether this effect could be mediated by an enhancement of the heat shock response. (2) Methods: Heat shock protein expression was analyzed in vitro and in vivo. Histamine was provided to primary microglia and NSC-34 motor neurons expressing the SOD1-G93A mutation. The brain permeable histamine precursor histidine was chronically administered to symptomatic SOD1-G93A mice. Spine density was measured by Golgi-staining in motor cortex of histidine-treated SOD1-G93A mice. (3) Results: We demonstrate that histamine activates the heat shock response in cultured SOD1-G93A microglia and motor neurons. In SOD1-G93A mice, histidine augments the protein content of GRP78 and Hsp70 in spinal cord and cortex, where the treatment also rescues type I motor neuron dendritic spine loss. (4) Conclusion: Besides the established histaminergic neuroprotective and anti-inflammatory effects, the induction of the heat shock response in the SOD1-G93A model by histamine confirms the importance of this pathway in the search for successful therapeutic solutions to treat ALS.

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Impaired proteostasis in senescent vascular endothelial cells: a perspective on estrogen and oxidative stress in the aging vasculature

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00318.2018 ◽

2019 ◽

Vol 316 (2) ◽

pp. H421-H429 ◽

Cited By ~ 3

Author(s):

HyunTae V. Hwang ◽

Yun Lin ◽

Michelle N. Rebuffatti ◽

Darlene T. Tran ◽

Lily Lee ◽

...

Keyword(s):

Oxidative Stress ◽

Endothelial Cells ◽

Heat Shock ◽

Protein Aggregation ◽

Cellular Senescence ◽

Heat Shock Response ◽

Human Endothelial Cells ◽

Early Passage ◽

Shock Response ◽

17Β Estradiol

The heat shock response is an important cytoprotective mechanism for protein homeostasis and is an essential protective response to cellular stress and injury. Studies on changes in the heat shock response with aging have been mixed with regard to whether it is inhibited, and this, at least in part, reflects different tissues and different models. Cellular senescence is a key feature in aging, but work on the heat shock response in cultured senescent (SEN) cells has largely been limited to fibroblasts. Given the prevalence of oxidative injury in the aging cardiovascular system, we investigated whether SEN primary human coronary artery endothelial cells have a diminished heat shock response and impaired proteostasis. In addition, we tested whether this downregulation of heat shock response can be mitigated by 17β-estradiol (E2), which has a critical cardioprotective role in women, as we have previously reported that E2 improves the heat shock response in endothelial cells (Hamilton KL, Mbai FN, Gupta S, Knowlton AA. Arterioscler Thromb Vasc Biol 24: 1628–1633, 2004). We found that SEN endothelial cells, despite their unexpectedly increased proteasome activity, had a diminished heat shock response and had more protein aggregation than early passage cells. SEN cells had increased oxidative stress, which promoted protein aggregation. E2 treatment did not decrease protein aggregation or improve the heat shock response in either early passage or SEN cells. In summary, cellular senescence in adult human endothelial cells is accompanied by increased oxidative stress and a blunting of proteostasis, and E2 did not mitigate these changes. NEW & NOTEWORTHY Senescent human endothelial cells have a diminished heat shock response and increased protein aggregates. Senescent human endothelial cells have increased basal oxidative stress, which increases protein aggregates. Physiological level of 17β-estradiol did not improve proteostasis in endothelial cells. Listen to this article's corresponding podcast at https://ajpheart.podbean.com/e/proteostasis-in-senescent-endothelial-cells/ .

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Unrestrained AMPylation targets cytosolic chaperones and activates the heat shock response

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1619234114 ◽

2016 ◽

Vol 114 (2) ◽

pp. E152-E160 ◽

Cited By ~ 21

Author(s):

Matthias C. Truttmann ◽

Xu Zheng ◽

Leo Hanke ◽

Jadyn R. Damon ◽

Monique Grootveld ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Heat Shock ◽

Protein Aggregation ◽

Heat Shock Response ◽

Protein Translation ◽

Cellular Protein ◽

Interacting Protein ◽

Endogenous Protein ◽

Shock Response ◽

Influenza Virus Replication

Protein AMPylation is a conserved posttranslational modification with emerging roles in endoplasmic reticulum homeostasis. However, the range of substrates and cell biological consequences of AMPylation remain poorly defined. We expressed human and Caenorhabditis elegans AMPylation enzymes—huntingtin yeast-interacting protein E (HYPE) and filamentation-induced by cyclic AMP (FIC)-1, respectively—in Saccharomyces cerevisiae, a eukaryote that lacks endogenous protein AMPylation. Expression of HYPE and FIC-1 in yeast induced a strong cytoplasmic Hsf1-mediated heat shock response, accompanied by attenuation of protein translation, massive protein aggregation, growth arrest, and lethality. Overexpression of Ssa2, a cytosolic heat shock protein (Hsp)70, was sufficient to partially rescue growth. In human cell lines, overexpression of active HYPE similarly induced protein aggregation and the HSF1-dependent heat shock response. Excessive AMPylation also abolished HSP70-dependent influenza virus replication. Our findings suggest a mode of Hsp70 inactivation by AMPylation and point toward a role for protein AMPylation in the regulation of cellular protein homeostasis beyond the endoplasmic reticulum.

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