Chromosomal mapping of core histone acetylation by immunoselection

Methods : a Companion to Methods in Enzymology
C Crane-RobinsonA W Thorne

Abstract

Acetylation of specific lysine residues in the N-terminal domains of core histones is a biochemical marker of active genes. To determine the spatial and temporal distribution of this reversible posttranslational modification, affinity-purified polyclonal antibodies recognizing the epitope epsilon-acetyllysine have been used in immunoselection procedures with mononucleosomes and salt-soluble chromatin fragments generated by micrococcal nuclease. The DNA of the antibody-selected chromatin was slot-blotted and probed with a variety of gene sequences: an enhanced hybridization signal, with respect to that from the DNA of the input chromatin, demonstrated elevated acetylation levels on the histones associated with the probing sequences. Using chicken embryonic erythrocytes as chromatin source and probes from the beta globin locus, it was shown that both the embryonic epsilon and adult beta genes are acetylated at 5 and 15 days, and the acetylation uniformly covers the whole of the locus, precisely comapping with the 33 kb of open chromatin structure. Studies with proliferating human K562 cells show that the inactive but poised PDGF-beta gene is already hyperacetylated and that its acetylation status is not enhanced on induction. The...Continue Reading

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Citations

Jul 21, 1998·Annals of the New York Academy of Sciences·G D GinderS Z Wang
Apr 13, 2000·The International Journal of Biochemistry & Cell Biology·L TorreM A Avila
Feb 12, 2009·Genome Biology·John Arne DahlPhilippe Collas
Mar 10, 2011·PloS One·Patrick O McGowanMoshe Szyf
May 27, 2016·Epigenetics & Chromatin·Sanzida JahanJames R Davie
Dec 22, 2006·PloS One·Alon GorenHoward Cedar
May 2, 2000·The Journal of Biological Chemistry·F ElefantS A Liebhaber

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