Chromatin position effects assayed by thousands of reporters integrated in parallel

Reporter genes integrated into the genome are a powerful tool to reveal effects of regulatory elements and local chromatin context on gene expression. However, so far such reporter assays have been of low throughput. Here, we describe a multiplexing approach for the parallel monitoring of transcript...

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Veröffentlicht in:Cell Jg. 154; H. 4; S. 914
Hauptverfasser: Akhtar, Waseem, de Jong, Johann, Pindyurin, Alexey V, Pagie, Ludo, Meuleman, Wouter, de Ridder, Jeroen, Berns, Anton, Wessels, Lodewyk F A, van Lohuizen, Maarten, van Steensel, Bas
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States 15.08.2013
Schlagworte:
Animals
Chromatin - metabolism
Chromosomal Position Effects
Embryonic Stem Cells - metabolism
Genes, Reporter
Genetic Techniques
High-Throughput Nucleotide Sequencing
Mice
Promoter Regions, Genetic
ISSN:1097-4172, 1097-4172
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Zusammenfassung:Reporter genes integrated into the genome are a powerful tool to reveal effects of regulatory elements and local chromatin context on gene expression. However, so far such reporter assays have been of low throughput. Here, we describe a multiplexing approach for the parallel monitoring of transcriptional activity of thousands of randomly integrated reporters. More than 27,000 distinct reporter integrations in mouse embryonic stem cells, obtained with two different promoters, show ∼1,000-fold variation in expression levels. Data analysis indicates that lamina-associated domains act as attenuators of transcription, likely by reducing access of transcription factors to binding sites. Furthermore, chromatin compaction is predictive of reporter activity. We also found evidence for crosstalk between neighboring genes and estimate that enhancers can influence gene expression on average over ∼20 kb. The multiplexed reporter assay is highly flexible in design and can be modified to query a wide range of aspects of gene regulation.
Bibliographie:ObjectType-Article-1
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ISSN:1097-4172
1097-4172
DOI:10.1016/j.cell.2013.07.018