Inferring transcriptional regulatory networks by integrative analysis in arabidopsis seed germination

Abstract

Living cells must continually adapt to changing environments by altering their gene expression patterns. One of the central regulatory mechanisms is transcriptional regulatory interactions between transcription factors and their target genes. There have been many experimental and computational approaches to solve this problem. As an experimental approach, microarray data have been applied to find a set of genes believed to be co-regulated with a transcription factor. However, many co-regulated genes can be indirectly affected by the transcription factor. To verify whether a target gene is regulated directly or not, ChIP-on-chip technology has been widely adopted. However, since ChIP-on-chip data provide us approximate binding sites in genome-wide scale, the motif discovery process using computational programs is further required to identify exact binding motifs. In this study, the transcriptional regulatory networks of Arabidopsis seed germination controlled by PIL5 were investigated by applying integrative analysis. PIL5 is a basic helix-loop-helix (bHLH) transcription factor that inhibits seed germination by regulating the expression of gibberellin (GA)- and Abscisic acid (ABA)-related genes either directly or indirectly. However, the comprehensive target gene networks and DNA-binding mechanisms are not yet known. First, total 748 novel PIL5 binding sites in the $\It{Arabidopsis}$ whole genome were identified using Chromatin immunoprecipitation (ChIP)-chip assay. Most of PIL5 binding sites were located in the promoter region consistent with its molecular function as transcription regulator. Gene expression profiling assay revealed that large numbers of gene expressions are regulated by PIL5 in response to light during seed germination. Comparison analysis of the ChIP-chip assay with the gene expressions profiling assay identified 166 PIL5 direct-regulated genes. Second, the in vivo binding motifs of PIL5 were identified by Oligo-analysis combined with K...