Architectural roles of multiple chromatin insulators at the human apolipoprotein gene cluster
Supplementary Material
Supplementary Figures S1–S8
Supplementary Table 1
Supplementary Information
Abstract
Long-range regulatory elements and higher-order chromatin structure coordinate the expression of multiple genes in cluster, and CTCF/cohesin-mediated chromatin insulator may be a key in this regulation. The human apolipoprotein (APO) A1/C3/A4/A5 gene region, whose alterations increase the risk of dyslipidemia and atherosclerosis, is partitioned at least by three CTCF-enriched sites and three cohesin protein RAD21-enriched sites (two overlap with the CTCF sites), resulting in the formation of two transcribed chromatin loops by interactions between insulators. The C3 enhancer and APOC3/A4/A5 promoters reside in the same loop, where the APOC3/A4 promoters are pointed towards the C3 enhancer, whereas the APOA1 promoter is present in the different loop. The depletion of either CTCF or RAD21 disrupts the chromatin loop structure, together with significant changes in the APO expression and the localization of transcription factor hepatocyte nuclear factor (HNF)-4α and transcriptionally active form of RNA polymerase II at the APO promoters. Thus, CTCF/cohesin-mediated insulators maintain the chromatin loop formation and the localization of transcriptional apparatus at the promoters, suggesting an essential role of chromatin insulation in controlling the expression of clustered genes.
Acknowledgments
We appreciate Dr Jan-Michael Peters (Research Institute of Molecular Pathology) for collaboration using ChIP-on-chip analyses, and the members in our laboratory for helpful discussions. This work was supported by a Grant-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Culture, Sports, Science and Technology (MN), by a Grant-in-Aid for the Global Center of Excellence (COE) ‘Cell Fate Regulation Research and Education Unit', Kumamoto University, and in part by a research grant from the Uehara Memorial Foundation (MN).