Fig. 1: A schematic depiction of the complex looping that facilitates transcriptional regulation and enables the coordinated regulation of multiple genes simultaneously. The looping is revealed by identifying chromosomal segments that interact throughout the genome (background).
© 2009 A*STAR
The textbook representation of a gene is deceptively simple: a line of DNA sequence punctuated by regulatory regions atop of which perch diverse activator and inhibitor proteins.
However, the reality within the nucleus is considerably more complex. Many of these regulatory sites reside surprisingly far from the transcriptional regions they govern, and evidence suggests that their activity may be dependent on the three-dimensional rearrangement of chromosomes to enable physical associations between spatially remote genomic segments.
“A few studies based on individual genes have suggested that long-distance elements may interact with gene promoters to regulate gene transcription,” explains Yijun Ruan of the A*STAR Genome Institute of Singapore. However, no method existed to study this theory at a genome-wide scale.
To address this challenge, Ruan and co-workers devised a technique called chromatin interaction analysis by paired-end tag sequencing (ChIA-PET), which enables global mapping of such interactions. Chromosomal DNA is normally wound around proteins to form material known as chromatin, and gene activation is typically accompanied by chemical modification and rearrangement of these proteins. ChIA-PET employs a strategy to physically link interacting chromatin regions; analysis of the DNA fragments contained within this chromatin subsequently reveals the specific genomic sites connected by these interactions (Fig. 1).
As an initial test, Ruan’s team performed ChIA-PET analysis of gene activation mediated by estrogen receptor α in human breast cancer cells. This revealed numerous chromatin loop structures connected at activated ‘anchor genes’, in which distal regulatory regions were brought into direct contact with gene promoters. In many cases, these individual duplexes were themselves part of larger, multi-loop arrangements, connecting multiple anchor genes while physically excluding other ‘loop genes’—and these interactions had a direct bearing on gene activity. “We [observed] that certain genes are looped together into the same transcription cluster for coordinated expression,” says Ruan. “We also provided evidence that relatively small loopings bring genes to the same transcription center for activation, while large loops may partition genes to inactive nuclear regions.”
These findings add an essential new dimension to genomic analysis, and Ruan is already planning experiments to extend ChIA-PET analysis to characterize activity of other key transcriptional activators, like the tumor suppressor p53. “Our study reveals that chromatin looping is a primary mechanism for coordinating gene transcription regulation,” he says, “[and] I envision that we may soon enter a ‘spatial genomics’ era, where we can view a genomic map in which regulatory elements interact with each other in three dimensions.”
The A*STAR-affiliated authors in this highlight are from the Genome Institute of Singapore.
