Rag2 accelerates lipofuscin accumulation in the brain: Implications for human stem cell brain transplantation studies.

Immunodeficient mice are widely used in human stem cell transplantation research. Recombination activating gene 1 (Rag1) deletion results in immunodeficiency and leads to accelerated aging in zebrafish with increased cytosolic accumulation of lipofuscin (LF). Unlike zebrafish, mammals have two homologs, Rag1 and Rag2, that regulate adaptive immunity.

Osmolytes dynamically regulate mutant Huntingtin aggregation and CREB function in Huntington's disease cell models.

Osmolytes are organic solutes that change the protein folding landscape shifting the equilibrium towards the folded state. Herein, we use osmolytes to probe the structuring and aggregation of the intrinsically disordered mutant Huntingtin (mHtt) vis-a-vis the pathogenicity of mHtt on transcription factor function and cell survival. Using an inducible PC12 cell model of Huntington's disease (HD), we show that stabilizing polyol osmolytes drive the aggregation of Htt103Q-EGFP from a diffuse ensemble into inclusion bodies (IBs), whereas the destabilizing osmolyte urea does not.

Heat shock promotes inclusion body formation of mutant huntingtin (mHtt) and alleviates mHtt-induced transcription factor dysfunction.

PolyQ-expanded huntingtin (mHtt) variants form aggregates, termed inclusion bodies (IBs), in individuals with and models of Huntington's disease (HD). The role of IB diffusible mHtt in neurotoxicity remains unclear. Using a ponasterone (PA)-inducible cell model of HD, here we evaluated the effects of heat shock on the appearance and functional outcome of Htt103Q-EGFP expression.

SIRT1 knockdown promotes neural differentiation and attenuates the heat shock response.

Neurons have a limited capacity for heat shock protein (HSP) induction and are vulnerable to the pathogenic consequence of protein misfolding and aggregation as seen in age-related neurodegenerative diseases. Sirtuin 1 (SIRT1), an NAD(+) -dependent lysine deacetylase with important biological functions, has been shown to sustain the DNA-binding state of HSF1 for HSP induction. Here we show that differentiation and maturation of embryonic cortical neurons and N2a neuroprogenitor cells is associated with decreases in SIRT1 expression and heat shock-dependent induction of HSP70 protein.

Glutamate dehydrogenase 1 and SIRT4 regulate glial development.

Congenital hyperinsulinism/hyperammonemia (HI/HA) syndrome is caused by an activation mutation of glutamate dehydrogenase 1 (GDH1), a mitochondrial enzyme responsible for the reversible interconversion between glutamate and α-ketoglutarate. The syndrome presents clinically with hyperammonemia, significant episodic hypoglycemia, seizures, and frequent incidences of developmental and learning defects. Clinical research has implicated that although some of the developmental and neurological defects may be attributed to hypoglycemia, some characteristics cannot be ascribed to low glucose and as hyperammonemia is generally mild and asymptomatic, there exists the possibility that altered GDH1 activity within the brain leads to some clinical changes.

Neuroprotective drug riluzole amplifies the heat shock factor 1 (HSF1)- and glutamate transporter 1 (GLT1)-dependent cytoprotective mechanisms for neuronal survival.

Heat shock factor 1 (HSF1) mediates the cellular response to stress to increase the production of heat shock protein (HSP) chaperones for proper protein folding, trafficking, and degradation; failure of this homeostatic mechanism likely contributes to neurodegeneration. We show that the neuroprotective drug riluzole increased the amount of HSF1 in NG108-15 neuroprogenitor cells by slowing the specific turnover of HSF1 and supporting a more robust and sustained activation of HSF1. Using Hsp70-luciferase as a functional readout of the activity of HSF1, we show that riluzole amplified the heat shock induction of the reporter gene with an optimal increase at 1 μM.

Thermal killing of human colon cancer cells is associated with the loss of eukaryotic initiation factor 5A.

Heat-induced cell death appears to be a cell-specific event. Chronic heat stress was lethal to human colon cancer cells (Caco-2, HT29, and HCT116), but not to normal diploid fibroblasts and other cancer cells (BJ-T, WI38, HeLa, ovarian 2008, WI38VA). Acute heat stress (45-51 degrees C, 30 min) caused cell death of colon cancer cells during recovery at physiological temperature.

Aberrant regulation and modification of heat shock factor 1 in senescent human diploid fibroblasts.

Induction of the heat shock response (HSR), determined by hsp70-luciferase reporter and HSP70 protein expression, is attenuated as a function of age of the IMR-90 human diploid fibroblasts. To better understand the underlying mechanism, we evaluated changes in the regulation and function of the HSF1 transcription factor. We show that the activation of HSF1 both in vivo and in vitro was decreased as a function of age, and this was attributable to a change in the regulation of HSF1 as the abundance of HSF1 protein and mRNA was unaffected.

Riluzole increases the amount of latent HSF1 for an amplified heat shock response and cytoprotection.

Background: Induction of the heat shock response (HSR) and increased expression of the heat shock proteins (HSPs) provide mechanisms to ensure proper protein folding, trafficking, and disposition. The importance of HSPs is underscored by the understanding that protein mis-folding and aggregation contribute centrally to the pathogenesis of neurodegenerative diseases.

Methodology/principal Findings: We used a cell-based hsp70-luciferease reporter gene assay system to identify agents that modulate the HSR and show here that clinically relevant concentrations of the FDA-approved ALS drug riluzole significantly increased the heat shock induction of hsp70-luciferse reporter gene.

Changes in the regulation of heat shock gene expression in neuronal cell differentiation.

Neuronal differentiation of the NG108-15 neuroblastoma-glioma hybrid cells is accompanied by a marked attenuation in the heat shock induction of the Hsp70-firefly luciferase reporter gene activity. Analysis of the amount and activation of heat shock factor 1, induction of mRNA(hsp), and the synthesis and accumulation of heat shock proteins (HSPs) in the undifferentiated and differentiated cells suggest a transcriptional mechanism for this attenuation. Concomitant with a decreased induction of the 72-kDa Hsp70 protein in the differentiated cells, there is an increased abundance of the constitutive 73-kDa Hsc70, a protein known to function in vesicle trafficking.

Neural differentiation and the attenuated heat shock response.

Differentiation of neural progenitor cells of neuroblastoma, pheochromocytoma, and surrogate stem cell lineages from a state resembling stem cells to a state resembling neurons is accompanied by a marked attenuation in induction of the heat shock protein 70 promoter driven-luciferase reporter gene, and induction of the reporter gene in primary embryonic neurons from hippocampus, cortex, and spinal cord is lower still when compared to the differentiated cells. Neural specificity of this phenotype is demonstrated by a negative correlation of hsp70-reporter gene expression and neurite extension under various experimental conditions. Analysis of biochemical events involved in induction of the heat shock response (HSR) reveal a blunted activation of HSF1 DNA-binding activity, and decreased induction of the mRNA(hsp70) and the 72 kDa HSP70 protein.

Dynamic regulation and involvement of the heat shock transcriptional response in arsenic carcinogenesis.

The objective of this study is to better define induction of the heat shock response by arsenite, and to evaluation if induction of heat shock proteins (HSPs) contributes to the carcinogenic activity of arsenite. We show here that arsenite is a ubiquitous inducer of the heat shock response in mammalian cells: that it activated heat shock transcription factor 1 (HSF1) DNA-binding activity, enhanced hsp 70 promoter, and induced hsp70mRNA and synthesis of HSP chaperones. Using a high throughput hsp70 promoter-luciferase reporter assay, we observed a hormetic dose response where low concentrations of arsenite stimulated and high concentrations inhibited.

Redox-dependent regulation of the conformation and function of human heat shock factor 1.

We present here evidence that redox-dependent thiol-disulfide exchange can provide a mechanism regulating the conformation and activity of the human heat shock transcription factor 1 (HSF1). Diamide and dithiothreitol were reagents used respectively to promote and reverse disulfide cross-link, and the resolution and detection of redox conformers of HSF1 were done according to recently published methods [Manalo, D. J.