The HumanGATA1Gene Retains a 5′ Insulator That Maintains Chromosomal Architecture andGATA1Expression Levels in Splenic Erythroblasts

Abstract

GATA1 is a key transcription factor for erythropoiesis.GATA1gene expression is strictly regulated at the transcriptional level. While the regulatory mechanisms governing mouseGata1(mGata1) gene expression have been studied extensively, how expression of the humanGATA1(hGATA1) gene is regulated remains to be elucidated. To address this issue, we generatedhGATA1bacterial artificial chromosome (BAC) transgenic mouse lines harboring a 183-kbhGATA1locus covering thehGATA1exons and distal flanking sequences. TransgenichGATA1expression coincides with endogenousmGata1expression and fully rescues hematopoietic deficiency inmGata1knockdown mice. The transgene exhibited copy number-dependent and integration position-independent expression ofhGATA1, indicating the presence of chromatin insulator activity within the transgene. We found a novel insulator element at 29 kb 5′ to thehGATA1gene and refer to this element as the 5′ CCCTC-binding factor (CTCF) site. Substitution mutation of the 5′ CTCF site in thehGATA1BAC disrupted the chromatin architecture and led to a reduction ofhGATA1expression in splenic erythroblasts under conditions of stress erythropoiesis. Our results demonstrate that expression of thehGATA1gene is regulated through the chromatin architecture organized by 5′ CTCF site-mediated intrachromosomal interactions in thehGATA1locus.