Genomic interaction profiles in breast cancer reveal altered chromatin architecture.

Gene transcription can be regulated by remote enhancer regions through chromosome looping either in cis or in trans. Cancer cells are characterized by wholesale changes in long-range gene interactions, but the role that these long-range interactions play in cancer progression and metastasis is not well understood. In this study, we used IGFBP3, a gene involved in breast cancer pathogenesis, as bait in a 4C-seq experiment comparing normal breast cells (HMEC) with two breast cancer cell lines (MCF7, an ER positive cell line, and MDA-MB-231, a triple negative cell line). The IGFBP3 long-range interaction profile was substantially altered in breast cancer. Many interactions seen in normal breast cells are lost and novel interactions appear in cancer lines. We found that in HMEC, the breast carcinoma amplified sequence gene family (BCAS) 1-4 were among the top 10 most significantly enriched regions of interaction with IGFBP3. 3D-FISH analysis indicated that the translocation-prone BCAS genes, which are located on chromosomes 1, 17, and 20, are in close physical proximity with IGFBP3 and each other in normal breast cells. We also found that epidermal growth factor receptor (EGFR), a gene implicated in tumorigenesis, interacts significantly with IGFBP3 and that this interaction may play a role in their regulation. Breakpoint analysis suggests that when an IGFBP3 interacting region undergoes a translocation an additional interaction detectable by 4C is gained. Overall, our data from multiple lines of evidence suggest an important role for long-range chromosomal interactions in the pathogenesis of cancer.

Interruption of intrachromosomal looping by CCCTC binding factor decoy proteins abrogates genomic imprinting of human insulin-like growth factor II.

Monoallelic expression of IGF2 is regulated by CCCTC binding factor (CTCF) binding to the imprinting control region (ICR) on the maternal allele, with subsequent formation of an intrachromosomal loop to the promoter region. The N-terminal domain of CTCF interacts with SUZ12, part of the polycomb repressive complex-2 (PRC2), to silence the maternal allele. We synthesized decoy CTCF proteins, fusing the CTCF deoxyribonucleic acid-binding zinc finger domain to CpG methyltransferase Sss1 or to enhanced green fluorescent protein. In normal human fibroblasts and breast cancer MCF7 cell lines, the CTCF decoy proteins bound to the unmethylated ICR and to the IGF2 promoter region but did not interact with SUZ12. EZH2, another part of PRC2, was unable to methylate histone H3-K27 in the IGF2 promoter region, resulting in reactivation of the imprinted allele. The intrachromosomal loop between the maternal ICR and the IGF2 promoters was not observed when IGF2 imprinting was lost. CTCF epigenetically governs allelic gene expression of IGF2 by orchestrating chromatin loop structures involving PRC2.

Chemokinetic effect of interleukin-1 beta on cultured Biomphalaria glabrata embryonic cells.

Previous studies have indicated that a molecule with cytokine activity, possibly an interleukin-1-like (IL-1) molecule, plays a role in the killing of larval stages of the blood fluke Schistosoma mansoni in the snail host Biomphalaria glabrata. The purpose of the present experiment was to test the effects of recombinant-human IL-1beta (rhIL-1beta) on embryonic B. glabrata (Bge) cell motility to determine whether the cells respond to the cytokine. Response was measured using a variation of a chemokinetic assay in which cells in culture were separated from variable concentrations of rhIL-1beta by a semi-permeable membrane containing pores to allow migration. A double staining technique was developed to ascertain cell movement across the membrane. The number of cells moving across the membrane significantly increased in a concentration-dependent manner relative to the presence of increasing amounts of rhIL-1beta below the membrane. The number of cells that moved across the membrane increased until a threshold was reached, after which migration decreased. Further, the rhIL-1beta-mediated increase in Bge cell migration across the membrane was abrogated by the addition of IL-1 receptor antagonist protein. These data indicate that Bge cells respond specifically to rhIL-1beta. As such, these data also indicate that Bge cells may serve as a useful model for elucidation of the role of cytokines or cytokine-like molecules in the snail/schistosome relationship.