Characterizing HSF1 Binding and Post-Translational Modifications of hsp70 Promoter in Cultured Cortical Neurons: Implications in the Heat-Shock Response.

Andrea V Gómez,Gonzalo Córdova,Gonzalo I Cancino,Alejandra R Álvarez,Michael Sherman,Álvaro P Barrios,Roberto Munita,Guillermo E Parada,María E Andrés

doi:10.1371/journal.pone.0129329

Andrea V Gómez, Gonzalo Córdova + Show 7 more

Open Access

https://doi.org/10.1371/journal.pone.0129329

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Journal: PloS one	Publication Date: Jun 8, 2015
Citations: 55	License type: CC BY 4.0

Affiliation: Pontificia Universidad Católica de Chile

Abstract

Causes of lower induction of Hsp70 in neurons during heat shock are still a matter of debate. To further inquire into the mechanisms regulating Hsp70 expression in neurons, we studied the activity of Heat Shock Factor 1 (HSF1) and histone posttranslational modifications (PTMs) at the hsp70 promoter in rat cortical neurons. Heat shock induced a transient and efficient translocation of HSF1 to neuronal nuclei. However, no binding of HSF1 at the hsp70 promoter was detected while it bound to the hsp25 promoter in cortical neurons during heat shock. Histone PTMs analysis showed that the hsp70 promoter harbors lower levels of histone H3 and H4 acetylation in cortical neurons compared to PC12 cells under basal conditions. Transcriptomic profiling data analysis showed a predominant usage of cryptic transcriptional start sites at hsp70 gene in the rat cerebral cortex, compared with the whole brain. These data support a weaker activation of hsp70 canonical promoter. Heat shock increased H3Ac at the hsp70 promoter in PC12 cells, which correlated with increased Hsp70 expression while no modifications occurred at the hsp70 promoter in cortical neurons. Increased histone H3 acetylation by Trichostatin A led to hsp70 mRNA and protein induction in cortical neurons. In conclusion, we found that two independent mechanisms maintain a lower induction of Hsp70 in cortical neurons. First, HSF1 fails to bind specifically to the hsp70 promoter in cortical neurons during heat shock and, second, the hsp70 promoter is less accessible in neurons compared to non-neuronal cells due to histone deacetylases repression.

Highlights

Heat, free radicals, bacterial infections, heavy metals, among other stresses, turn on the heat shock response in cells
The amount of Hsp70 protein increased in the three cell types in response to heat shock, even though the data confirm that the increment of Hsp70 in cells with neuronal phenotype, such as neuronal growth factor (NGF)-treated PC12 cells (PC12 plus NGF, 3.9 ± 0.3 fold of induction) and cortical neurons (2.4 ± 0.6 fold of induction) is significantly lower than in non-neuronal cells (Fig 1A)
After 2h of heat shock, the mRNA of hsp70 is induced 96.8 times in undifferentiated PC12 cells while in cortical neurons is induced only 8.9 times (Fig 1B). This effect is specific for hsp70 gene since hsp25 mRNA is induced to a similar extent in cortical neurons and, NGF-treated and naïve PC12 cells (Fig 1C)

Summary

Introduction

Free radicals, bacterial infections, heavy metals, among other stresses, turn on the heat shock response in cells. Guertin and Lis [24] showed in Drosophila by genome-wide analysis that an active chromatin landscape around HSEs is required for HSF1 binding elicited by heat shock. These data indicate that the regulation of Hsp expression and neuroprotection mechanisms during stress in neurons are still poorly understood. The importance of PTMs of histones on hsp promoter in response to stress has been shown in yeast and Drosophila and increasing data is available in mammalian genomes [26]. The data show that cortical neurons display lower response to heat shock even though HSF1 is present and activated. Strong HDAC-dependent repression and a specific failure of HSF1 binding to the hsp promoter weakens Hsp induction in cortical neurons

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Discussion