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.


RESULTS
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. Immuno-Western and -cytochemical analysis of HSF1 show that riluzole increased the amount of cytosolic HSF1 to afford a greater activation of HSF1 upon heat shock. The increased HSF1 contributed centrally to the cytoprotective activity of riluzole as hsf1 gene knockout negated the synergistic activity of riluzole and conditioning heat shock to confer cell survival under oxidative stress. Evidence of a post-transcriptional mechanism for the increase in HSF1 include: quantitation of mRNA(hsf1) by RT-PCR showed no effect of either heat shock or riluzole treatment; riluzole also increased the expression of HSF1 from a CMV-promoter; analysis of the turnover of HSF1 by pulse chase and immunoprecipitation show that riluzole slowed the decay of [(35)S]labeled-HSF1. The effect of riluzole on HSF1 was qualitatively different from that of MG132 and chloroquine, inhibitors of the proteasome and lysosome, respectively, and appeared to involve the chaperone-mediated autophagy pathway as RNAi-mediated knockdown of CMA negated its effect.


CONCLUSIONS
We show that riluzole increased the amount of HSF1 to amplify the HSR for cytoprotection. Our study provides novel insight into the mechanism that regulates HSF1 turnover, and identifies the degradation of HSF1 as a target for therapeutics intervention.
Yang
Jingxian
J
Department of Cell Biology and Neuroscience, Rutgers State University of New Jersey, Piscataway, New Jersey, United States of America.
Bridges
Kristen
K
Chen
Kuang Yu
KY
Liu
Alice Y-C
AY
D002470:Cell Survival / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics*; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D017930:Genes, Reporter; D006367:HeLa Cells; D000076249:Heat Shock Transcription Factors; D006358:Hot Temperature; D006801:Humans; D008156:Luciferases / Q000235:genetics; D019636:Neurodegenerative Diseases / Q000188:drug therapy / Q000503:physiopathology; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D019782:Riluzole / Q000494:pharmacology*; D013116:Spinal Cord / Q000196:embryology / Q000502:physiology; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics*
D004268:DNA-Binding Proteins; D018690:Excitatory Amino Acid Agonists; C000619706:HSF1 protein, human; D000076249:Heat Shock Transcription Factors; D014157:Transcription Factors; D019782:Riluzole; D008156:Luciferases