title: Evaluation of Innate Immune Gene Expression Following HDAC Inhibitor Treatment by High Throughput qPCR and PhosFlow Cytometry.
abstract: The dynamics of chromatin structure contribute to the regulation of gene transcription and in part, the changes in chromatin structure associated with gene activation/repression are a function of the state of histone acetylation. Histone deacetylases (HDACs) deacetylate histone tails leading to a more compact structure of chromatin that in turn represses gene transcription. Given the rapid activation and/or repression of gene networks following microbial infection, the role of HDACs in the epigenetic regulation of genes involved in the innate and adaptive immune responses has become an area of extensive research. In relation to the immune-modulatory properties of HDAC inhibitors, we provide in the following methodological article an extended description of two techniques-a high throughput qPCR assay combined with PhosFlow cytometry-to evaluate the modulation of antiviral and inflammatory signaling cascades following HDAC inhibitor treatment. The high-throughput qPCR assay is based on the nanofluidic Fluidigm BioMark system that permits the analysis of up to 9216 qPCR reactions at once in a self-design open array chip. Together with the more refined analysis provided with the Phosflow technique, these two strategies offer invaluable tools to measure modulation of innate immune gene networks.
authors.affiliation: Lady Davis Institute-Jewish General Hospital, McGill University, Montreal, QC, Canada.
authors.forename: David
authors.initials: D
authors.lastname: Olagnier
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Sapienza University, Rome, Italy.
authors.forename: Cindy
authors.initials: C
authors.lastname: Chiang
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Sapienza University, Rome, Italy. john.hiscott@istitutopasteur.it.
authors.forename: John
authors.initials: J
authors.lastname: Hiscott
mesh_terms: D000107:Acetylation; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects* / Q000276:immunology; D044127:Epigenesis, Genetic; D005434:Flow Cytometry / Q000379:methods*; D057166:High-Throughput Screening Assays; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology*; D006655:Histone Deacetylases / Q000235:genetics* / Q000276:immunology; D006657:Histones / Q000235:genetics / Q000276:immunology; D006801:Humans; D007113:Immunity, Innate; D007371:Interferon-gamma / Q000494:pharmacology; D016328:NF-kappa B / Q000235:genetics / Q000276:immunology; D010766:Phosphorylation; D060888:Real-Time Polymerase Chain Reaction; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000276:immunology; D014158:Transcription, Genetic
chemical_list: D002843:Chromatin; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D016328:NF-kappa B; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D007371:Interferon-gamma; D006655:Histone Deacetylases
keywords: Antiviral response; Cytometry; Gene transcription; HDAC; HDAC inhibitor; High throughput qPCR; Immune response; Phosflow