Profiling histone modification patterns in plants using genomic tiling microarrays

In eukaryotes, the processes of transcription, replication and homologous recombination occur in the context of chromatin. Although tightly packed, this structure is highly dynamic, being modified through the action of many enzymatic activities that reorganize nucleosomes; covalently modify histones through acetylation, phosphorylation, methylation and so on; exchange histones with variants; and in many eukaryotes, including plants and mammals, methylate DNA residues(1). Transcriptional activity is usually associated with hyperacetylation of histones tails as well as with di- or trimethylation of Lys4 in histone H3. Conversely, silent chromatin typically correlates with histone hypoacetylation and di- or trimethylation of histone H3 Lys9 (refs. 2,3). Because of its small genome, Arabidopsis thaliana serves as a powerful system for understanding the role of various histone modifications in a complex organism, notably in association with DNA methylation and in relation to the epigenetic inheritance of silent chromatin(4). Here we describe a chromatin immunoprecipitation (ChIP) protocol developed for A. thaliana that permits, in combination with hybridization to genomic tiling microarrays, the mapping of histone modifications with high resolution along large genomic regions(5). After cross-linking, chromatin is immunoprecipitated using antibodies directed against specific histone modifications. DNA recovered from the precipitate is amplified, Labeled, hybridized to microarrays and compared to total DNA. This protocol has been used successfully to map histone H3 methylated at Lys4 or Lys9 across a 1.5-Mb region(5) and should have broad applications in plants and other organisms. A protocol that outlines the profiling of DNA methylation patterns at similar high resolution has also been developed(6).