PARP1 enhances inflammatory cytokine expression by alteration of promoter chromatin structure in microglia

Brain and Behavior

Ricardo Iván Martínez-Zamudio¹ 

[1] Department of Biochemistry and Molecular & Cellular Biology, Washington, District of Columbia

关键词: ADP‐ribosylation;    nucleosome;    inflammation;    gene expression;    cytokine;   

DOI  :  10.1002/brb3.239

来源: Wiley

【 摘 要 】

Abstract

Background

Poly(ADP-ribose) polymerase 1 (PARP1) is a chromatin-associated enzyme that participates in processes such as transcription and DNA repair through the regulation of chromatin structure. Accumulating evidence suggests an important role for PARP1 enzymatic activity in promoting CNS inflammation by facilitating the expression of inflammatory cytokines in glial cells. However, the molecular mechanisms by which PARP1 enzymatic activity mediates this process are not well understood. In this report we sought to determine the molecular mechanisms by which PARP1 enzymatic activity facilitates the expression of Il1β and TNF in LPS-stimulated BV2 cells.

Methods

PARP1 enzymatic activity and histone ADP-ribosylation were measured in LPS-stimulated BV2 cells by radioactive labelling with ³²P-NAD⁺. To assess the effect of histone ADP-ribosylation on nucleosome structure, in vitro nucleosome remodeling, nuclease accessibility and binding assays were performed. These studies were complemented by chromatin immunoprecipitation assays in resting and LPS-stimulated BV2 cells in order to determine the occupancy of PARP1, nucleosomes and the RelA subunit of NF-κB, as well as ADP-ribosylation, at the Il1β and Tnf promoters. Finally, we determined the effect of pharmacological inhibition of PARP1 enzymatic activity on the LPS stimulation-dependent induction of Il1β and Tnf mRNA.

Results

Our results indicate that LPS stimulation induces PARP1 enzymatic activity and histone ADP-ribosylation in the chromatin compartment of BV2 cells. In vitro studies show that nucleosome-bound PARP1 disrupts nucleosome structure histone ADP-ribosylation, increasing the accessibility of nucleosomal DNA. Consistent with this PARP1 is constitutively associated with at the Il1β and Tnf promoters in resting BV2 cells. Upon stimulation with LPS, ADP-ribosylation is observed at these promoters, and this is correlated with increased recruitment of the transcription factor NF-κB, resulting in robust transcription of these inflammatory cytokines. Accordingly, pharmacological inhibition of PARP1 enzymatic activity reduces NF-κB recruitment, and Il1β and Tnf expression in LPS-stimulated microglia.

Conclusions

Collectively, our data suggest that PARP1 facilitates inflammatory cytokine expression in microglia by increasing the accessibility of promoter DNA via histone ADP-riboyslation.

【 授权许可】

CC BY   
© 2014 The Authors. Brain and Behavior published by Wiley Periodicals, Inc.

Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.