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High-throughput screening for TLR3-IFN regulatory factor 3 signaling pathway modulators identifies several antipsychotic drugs as TLR inhibitors.

ABSTRACT: TLR3 is one of the major innate immune sensors of dsRNA. The signal transduction pathway activated by TLR3, upon binding to dsRNA, leads to the activation of two major transcription factors: NF-kappaB and IFN regulatory factor (IRF) 3. In an effort to identify specific chemical modulators of TLR3-IRF3 signal transduction pathway, we developed a cell-based readout system. Using the IFN-stimulated gene 56 promoter-driven firefly luciferase gene stably integrated in a TLR3-expressing HEK293 cell line, we were able to generate a cell line where treatment with dsRNA resulted in a dose-dependent induction of luciferase activity. A screen of two pharmacologically active compound libraries using this system identified a number of TLR3-IRF3 signaling pathway modulators. Among them we focused on a subset of inhibitors and characterized their mode of action. Several antipsychotic drugs, such as sertraline, trifluoperazine, and fluphenazine, were found to be direct inhibitors of the innate immune signaling pathway. These inhibitors also showed the ability to inhibit IFN-stimulated gene 56 induction mediated by TLR4 and TLR7/8 pathways. Interestingly, they did not show significant effects on TLR3-, TLR7-, and TLR8-mediated NF-kappaB activation. Detailed analysis of the signaling pathway indicated that these drugs might be exerting their inhibitory effects on IRF3 via PI3K signaling pathway. The data presented in this study provide mechanistic explanation of possible anti-inflammatory roles of some antipsychotic drugs.

SUBMITTER: Zhu J

PROVIDER: S-EPMC2874113 | biostudies-literature | 2010 May

REPOSITORIES: biostudies-literature

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High-throughput screening for TLR3-IFN regulatory factor 3 signaling pathway modulators identifies several antipsychotic drugs as TLR inhibitors.

Zhu Jianzhong J Smith Kevin K Hsieh Paishiun N PN Mburu Yvonne K YK Chattopadhyay Saurabh S Sen Ganes C GC Sarkar Saumendra N SN

Journal of immunology (Baltimore, Md. : 1950) 20100409 10

TLR3 is one of the major innate immune sensors of dsRNA. The signal transduction pathway activated by TLR3, upon binding to dsRNA, leads to the activation of two major transcription factors: NF-kappaB and IFN regulatory factor (IRF) 3. In an effort to identify specific chemical modulators of TLR3-IRF3 signal transduction pathway, we developed a cell-based readout system. Using the IFN-stimulated gene 56 promoter-driven firefly luciferase gene stably integrated in a TLR3-expressing HEK293 cell li ...[more]

PMID: 20382888

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Project description:BACKGROUND: Previous studies from our laboratory and others have demonstrated that in addition to altering chromatin acetylation and conformation, histone deacetylase inhibitors (HDACi) disrupt the acetylation status of numerous transcription factors and other proteins. A whole genome yeast deletion library screen was used to identify components of the transcriptional apparatus that modulate the sensitivity to the hydroxamic acid-based HDACi, CG-1521. RESULTS: Screening 4852 haploid Saccharomyces cerevisiae deletion strains for sensitivity to CG-1521 identifies 407 sensitive and 80 resistant strains. Gene ontology (GO) enrichment analysis shows that strains sensitive to CG-1521 are highly enriched in processes regulating chromatin remodeling and transcription as well as other ontologies, including vacuolar acidification and vesicle-mediated transport. CG-1521-resistant strains include those deficient in the regulation of transcription and tRNA modification. Components of the SAGA histone acetyltransferase (HAT) complex are overrepresented in the sensitive strains, including the catalytic subunit, Gcn5. Cell cycle analysis indicates that both the wild-type and gcn5? strains show a G1 delay after CG-1521 treatment, however the gcn5? strain displays increased sensitivity to CG-1521-induced cell death compared to the wild-type strain. To test whether the enzymatic activity of Gcn5 is necessary in the response to CG-1521, growth assays with a yeast strain expressing a catalytically inactive variant of the Gcn5 protein were performed and the results show that this strain is less sensitive to CG-1521 than the gcn5? strain. CONCLUSION: Genome-wide deletion mutant screening identifies biological processes that affect the sensitivity to the HDAC inhibitor CG-1521, including transcription and chromatin remodeling. This study illuminates the pathways involved in the response to CG-1521 in yeast and provides incentives to understand the mechanisms of HDAC inhibitors in cancer cells. The data presented here demonstrate that components of the SAGA complex are involved in mediating the response to CG-1521. Additional experiments suggest that functions other than the acetyltransferase activity of Gcn5 may be sufficient to attenuate the effects of CG-1521 on cell growth.

Dataset Information

High-throughput screening for TLR3-IFN regulatory factor 3 signaling pathway modulators identifies several antipsychotic drugs as TLR inhibitors.

Publications

High-throughput screening for TLR3-IFN regulatory factor 3 signaling pathway modulators identifies several antipsychotic drugs as TLR inhibitors.

Similar Datasets

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