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Dissecting primary (translation independent) from secondary (translation dependent) IFN-mediated differential gene expression

ABSTRACT: NIH-3T3 cells were pretreated for 15 min with either DMSO (mock) or cycloheximide followed by addition of either mock, 100 U/ml IFNalpha or 100 U/ml IFNgamma for 1h. During the last 30 min, 500 µM 4-thiouridine was added to cell culture medium. Total cellular RNA was isolated using Trizol reagent and nascent RNA was purified as described (Dölken et al. RNA 2008) . Three replicates of nascent RNA were analyzed by Affymetrix Mouse Gene ST 1.0 arrays Primary (translation independent) from secondary (translation dependent) IFN-mediated differential gene expression were studied in NIH-3T3 fibroblasts by studying studying differential gene expression in presence and absence of Cycloheximide (CHX). In addition, the effect of 75 min CHX during the last 30 min of treatment was studied in nascent RNA.

ORGANISM(S): Mus musculus

SUBMITTER: Lars Dölken 

PROVIDER: E-GEOD-30457 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Deciphering the modulation of gene expression by type I and II interferons combining 4sU-tagging, translational arrest and in silico promoter analysis.

Trilling Mirko M   Bellora Nicolás N   Rutkowski Andrzej J AJ   de Graaf Miranda M   Dickinson Paul P   Robertson Kevin K   Prazeres da Costa Olivia O   Ghazal Peter P   Friedel Caroline C CC   Albà M Mar MM   Dölken Lars L  

Nucleic acids research 20130705 17

Interferons (IFN) play a pivotal role in innate immunity, orchestrating a cell-intrinsic anti-pathogenic state and stimulating adaptive immune responses. The complex interplay between the primary response to IFNs and its modulation by positive and negative feedback loops is incompletely understood. Here, we implement the combination of high-resolution gene-expression profiling of nascent RNA with translational inhibition of secondary feedback by cycloheximide. Unexpectedly, this approach reveale  ...[more]

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