Whole Genome Chromatin Interaction Analysis of Transcriptional Regulation Networks in Human Cancer Cells
Authors: Yijun Ruan
Journal: Clinical Cancer Research (2007)

PL01-01  Whole Genome Analysis of Transcriptional Regulation Networks in Human Cancer Cells Using Paired End diTag (PET) Technology  Yijun Ruan, PhD  Genome Institute of Singapore  60 Biopolis Street, Singapore 138672  Modern biology research is largely and will be continuously relying on our ability to sequence and characterize the human genome and other model genomes. Rapid developments of next-generation sequencing technology and novel application strategies are evident in recent years. Our approach of making contribution is to improve tag-based sequencing strategy. We first developed the paired-end ditagging (PET) approach and devised the Gene Identification Signature (GIS-PET) analysis for accurate demarcation of boundaries of full-length transcripts (Ng et al. 2005 Nature Methods 2:105) and highly efficient identification of unconventional fusion transcripts in human cancer cells (Ruan et al. 2007 Genome Res 17:828). PET-based transcriptome/genome analysis is further enhanced by ‘short read’ but ‘highly paralleled” next-generation sequencing instruments (454/Roche, Solexa/Illumina, and SOLiD/ABI) for multiplex sequencing of paired-end ditags (MS-PET) (Ng et al. 2006 Nucleic Acid Research 34:e84). To identify and study promoters and regulatory elements that mediate gene transcription, we invented the ChIP-PET analysis (chromatin immuno-precipitation coupled with paired-end ditagging) for highly accurate, robust and unbiased genome-wide identification of transcription factor binding sites (Wei et al. 2006 Cell 124:207; Loh et al. 2006 Nature Genetics 38:431; Zeller et al. 2006 PNAS 103:17834; Lin et al. 2007 PLoS Genetics 3:e87; Lim et al. 2007 Mol Cell 27:622). A growing body of data has shown that a large portion of regulatory elements are localized far away from gene coding regions. This observation cannot be explained by a simple linear relationship along the genome, and our current technologies are inefficient to address non-linear interactions between functional DNA elements. To elucidate potential 3-dimentional interaction relationship of functional elements, we have developed a new strategy for whole genome Chromatin Interaction Analysis using Paired End diTagging (ChIA-PET). In this approach, long range chromatin and inter-chromosomal interactions mediated by protein factors are fixed and the fragmented DNA-protein complexes are enriched by chromatin immuno-precipitation. The tethered DNA fragments in each complex are jointed with linker sequence by ligation, and the ligation products are further analyzed by MS-PET sequencing. Consequently, the ChIA-PET sequences are mapped to the reference genome to reveal the relationship of remote chromosomal regions brought in close proximity by protein factors. Our results suggest that long-range and multiplex looping interaction is an extensive mechanism in regulating gene transcription. Together, PET-based DNA sequencing analyses have been enabling us to generate large volume data efficiently from various dimensions around transcription activity. The accumulation and integration of these datasets are starting to emerge a comprehensive picture of transcription regulation networks.