Amigo, J. D.Yu, M.Troadec, M.-B.Gwynn, B.Cooney, J. D.Lambert, A. J.Chi, N. C.Weiss, M. J.Peters, L. L.Kaplan, J.Cantor, AlanPaw, Barry Htin2015-07-062011Amigo, J. D., M. Yu, M.-B. Troadec, B. Gwynn, J. D. Cooney, A. J. Lambert, N. C. Chi, et al. 2011. “Identification of Distal Cis-Regulatory Elements at Mouse Mitoferrin Loci Using Zebrafish Transgenesis.” Molecular and Cellular Biology 31 (7) (January 19): 1344–1356. doi:10.1128/mcb.01010-10.0270-7306http://nrs.harvard.edu/urn-3:HUL.InstRepos:17085600Mitoferrin 1 (Mfrn1; Slc25a37) and mitoferrin 2 (Mfrn2; Slc25a28) function as essential mitochondrial iron importers for heme and Fe/S cluster biogenesis. A genetic deficiency of Mfrn1 results in a profound hypochromic anemia in vertebrate species. To map the cis-regulatory modules (CRMs) that control expression of the Mfrn genes, we utilized genome-wide chromatin immunoprecipitation (ChIP) datasets for the major erythroid transcription factor GATA-1. We identified the CRMs that faithfully drive the expression of Mfrn1 during blood and heart development and Mfrn2 ubiquitously. Through in vivo analyses of the Mfrn-CRMs in zebrafish and mouse, we demonstrate their functional and evolutionary conservation. Using knockdowns with morpholinos and cell sorting analysis in transgenic zebrafish embryos, we show that GATA-1 directly regulates the expression of Mfrn1. Mutagenesis of individual GATA-1 binding cis elements (GBE) demonstrated that at least two of the three GBE within this CRM are functionally required for GATA-mediated transcription of Mfrn1. Furthermore, ChIP assays demonstrate switching from GATA-2 to GATA-1 at these elements during erythroid maturation. Our results provide new insights into the genetic regulation of mitochondrial function and iron homeostasis and, more generally, illustrate the utility of genome-wide ChIP analysis combined with zebrafish transgenesis for identifying long-range transcriptional enhancers that regulate tissue development.en-USJournal Article2015-07-0610.1128/MCB.01010-10