Chromatin Immunoprecipitation to Identify Global Targets of WRKY Transcription Factor Family Members Involved in Plant Immunity

Abstract

The completion of the alfalfa, Arabidopsis, papaya, poplar, and rice genome sequences along with ongoing sequencing projects of various crop species, offers an excellent opportunity to study gene expression at the whole genome level and to unravel the complexity of gene networks underlying the reprogramming of plant defense toward pathogen challenge. Gene expression in eukaryotic cells is mainly controlled by regulatory elements that recruit transcription factors (TFs) to modulate transcriptional outputs. Therefore, methods allowing the identification of all cognate TF binding sites (TFBS) within the regulatory regions of target genes on a genome-wide basis are the next obvious step to elucidate the plant defense transcriptome. Chromatin immunoprecipitation (ChIP) is one such powerful technique for analyzing functional cis-regulatory DNA elements. The ChIP assay allows the identification of specific regulatory DNA regions associated with trans-acting regulatory factors in vivo. ChIP assays can provide spatial and temporal snapshots of the regulatory components involved in reprogramming host gene expression upon pathogen ingress. Moreover, the use of ChIP-enriched DNA for hybridization to tiling microarrays (ChIP-chip) or for direct sequencing (ChIP-Seq) by means of massively parallel sequencing technology has expanded this methodology to address global changes in gene expression.