Chromosome Conformation Capture Carbon Copy (5C): A massively parallel solution for mapping interactions between genomic elements

Abstract

Physical interactions between genetic elements located throughout the genome play important roles in gene regulation and can be identified with the Chromosome Conformation Capture (3C) methodology. 3C converts physical chromatin interactions into specific ligation products, which are quantified individually by PCR. Here we present a high-throughput 3C approach, 3C-Carbon Copy (5C), that employs microarrays or quantitative DNA sequencing using 454-technology as detection methods. We applied 5C to analyze a 400-kb region containing the human ␤-globin locus and a 100-kb conserved gene desert region. We validated 5C by detection of several previously identified looping interactions in the ␤-globin locus. We also identified a new looping interaction in K562 cells between the ␤-globin Locus Control Region and the ␥-␦-globin intergenic region. Interestingly, this region has been implicated in the control of developmental globin gene switching. 5C should be widely applicable for large-scale mapping of cis-and trans-interaction networks of genomic elements and for the study of higher-order chromosome structure.

Caveats in the Established Understanding of CMT1A

“…For instance, the duplicated 1.4 Mb DNA segment of chromosome 17p12 (c17p12) may have disrupted the normal genome structure. Intergene or interchromosomal interaction has been demonstrated to regulate genomic structure and expression of genes 26, 27. Normal trans interactions between two alleles of c17p12 (interchromatid) are likely altered by introducing an additional copy of c17p12 in CMT1A.…”

“…The chromosomal structure derived from the trans/intrachromosomal interactions has been achieved using C3 technique 28. More advanced versions of the technique (C4 or C5) are now also available 26, 27. To determine if the dynamic changes of the duplicated c17p12 structure are pathogenic, single cell clones (such as fibroblasts from human skin biopsy or immature Schwann cells by iPSC technology) may be isolated from humans with CMT1A.…”

Getting connected in the globin interactome

“…These loci are all biologically plausible candidates that are involved in pathways known to be important in lung biology, including immune function, muscle function or inflammation. These studies did not replicate the SERPINE2 association and also did not find evidence for association to the 15q CHRNA5-CHRNA3-CHRNB4 locus previously associated with nicotine dependence, lung cancer and COPD [11][12][13] . …”

Repeated elements coordinate the spatial organization of the yeast genome

“…It is clear that the spatial arrangement of the genome into subcompartments is important for the regulation of developmental and cellular processes (Chakalova et al, 2005;Dostie et al, 2006;Foster and Bridger, 2005;Lanctot et al, 2007;Ling et al, 2006;Spilianakis et al, 2005;Zhao et al, 2006). However, we currently do not know how nuclear subcompartments are formed.…”

“…This potential is exemplified by the fact that the interacting DNA sequences are often not linearly arranged or even on the same chromosome (Dostie et al, 2006;Ling et al, 2006;O'Sullivan et al, 2004;Simonis et al, 2006;Spilianakis et al, 2005;Zhao et al, 2006). Therefore, the formation of intra-or interchromosomal interactions specifies the relative positions of chromosomal sequences, due to the contiguous nature of the chromosome(s).…”