Quantitative differences in chromatin accessibility across regulatory regions can be directly compared in distinct cell-types.
Transcriptional activation in eukaryotes is often accompanied by alterations to chromatin structure at specific regulatory sites while other genomic regions may remain unchanged. In this study, we have examined the correlation between expression and chromatin accessibility of the human CR2 gene in a panel of cell lines (U937, REH, Ramos, and Raji) using the CHART-PCR assay with the accessibility agent micrococcal nuclease (MNase). To validate the use of this assay for comparing multiple cell-types, we first tested a series of genomic regions to determine if we could observe consistent, site-specific levels of MNase chromatin accessibility. Promoter regions of the ubiquitously expressed genes GAPDH and beta-actin were similar and showed high accessibility to MNase digestion in each of the cell lines, while on the other hand, promoter regions of developmentally restricted genes PAX-7 and SP-A2 showed consistently reduced chromatin accessibility. Since CHART-PCR detected site-specific differences in chromatin accessibility in a manner that could be compared between cell-types, we next examined chromatin accessibility over the CR2 core promoter in the panel of cell lines representing either CR2 expressing or CR2 non-expressing cell-types. Our data revealed significantly enhanced accessibility over the -289 to -101 and the -115 to -12 regions of the CR2 promoter in expressing B-cells (Ramos, Raji) compared to non-expressing cells (U937, REH). Thus, CHART-PCR assays detected a correlation between chromatin accessibility and expression of the human CR2 gene, while the accessibility of other genomic regions was site-specific, but not altered between cell-types.
Cruickshank
Mark
M
Biochemistry and Molecular Biology, School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia, 35 Stirling Highway, Crawley, 6009 WA, Australia.
Fenwick
Emily
E
Abraham
Lawrence J
LJ
Ulgiati
Daniela
D
D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000235:genetics*; D042002:Chromatin Assembly and Disassembly / Q000235:genetics*; D006801:Humans; D017464:Receptors, Complement 3d / Q000235:genetics*; D050436:Regulatory Elements, Transcriptional / Q000235:genetics*; D013045:Species Specificity; D015533:Transcriptional Activation / Q000235:genetics*
D002843:Chromatin; D017464:Receptors, Complement 3d