Abstract Recent evidence suggests that long-range enhancers and gene promoters are in close proximity, which might reflect the formation of chromatin loops. Here, we examined the mechanism for DNA looping at the β-globin locus. By using chromosome conformation capture (3C), we show that the hematopoietic transcription factor GATA-1 and its cofactor FOG-1 are required for the physical interaction between the β-globin locus control region (LCR) and the β-major globin promoter. Kinetic studies reveal that GATA-1-induced loop formation correlates with the onset of β-globin transcription and occurs independently of new protein synthesis. GATA-1 occupies the β-major globin promoter normally in fetal liver erythroblasts from mice lacking the LCR, suggesting that GATA-1 binding to the promoter and LCR are independent events that occur prior to loop formation. Together, these data demonstrate that GATA-1 and FOG-1 are essential anchors for a tissue-specific chromatin loop, providing general insights into long-range enhancer function.