RT Journal Article SR Electronic T1 Locus-specific ChIP combined with NGS analysis reveals genomic regulatory regions that physically interact with the Pax5 promoter in a chicken B cell line JF bioRxiv FD Cold Spring Harbor Laboratory SP 089821 DO 10.1101/089821 A1 Toshitsugu Fujita A1 Fusako Kitaura A1 Miyuki Yuno A1 Yutaka Suzuki A1 Sumio Sugano A1 Hodaka Fujii YR 2016 UL http://biorxiv.org/content/early/2016/11/26/089821.abstract AB Chromosomal interactions regulate genome functions, such as transcription, via dynamic chromosomal organization in the nucleus. In this study, we identified genomic regions that physically bind to the promoter region of the Pax5 gene in the chicken B-cell line DT40, with the goal of obtaining mechanistic insight into transcriptional regulation through chromosomal interaction. Using insertional chromatin immunoprecipitation (iChIP) in combination with next-generation sequencing (NGS) (iChIP-Seq), we found that the Pax5 promoter bound to multiple genomic regions. The identified chromosomal interactions were independently confirmed by in vitro engineered DNA-binding molecule-mediated ChIP (in vitro enChIP) in combination with NGS (in vitro enChIP-Seq). Comparing chromosomal interactions in wild-type DT40 with those in a macrophage-like counterpart, we found that some of the identified chromosomal interactions were organized in a B cell–specific manner. In addition, deletion of a B cell–specific interacting genomic region in chromosome 11, which was marked by active enhancer histone modifications, resulted in moderate but significant down-regulation of Pax5 transcription. Together, these results suggested that Pax5 transcription in DT40 cells is regulated by inter-chromosomal interactions. Moreover, these analyses showed that iChIP-Seq and in vitro enChIP-Seq are useful for non-biased identification of functional genomic regions that physically interact with a locus of interest.