The SCL +40 Enhancer Targets the Midbrain Together with Primitive and Definitive Hematopoiesis and Is Regulated by SCL and GATA Proteins

Abstract

The SCL/Tal-1 gene encodes a basic helix-loop-helix transcription factor with key roles in hematopoietic and neural development. SCL is expressed in, and required for, both primitive and definitive erythropoiesis. Thus far, we have identified only one erythroid SCL enhancer. Located 40 kb downstream of exon 1a, the +40 enhancer displays activity in primitive erythroblasts. We demonstrate here that a 3.7-kb fragment containing this element also targets expression to the midbrain, a known site of endogenous SCL expression. Although the 3.7-kb construct was active in primitive, but not definitive, erythroblasts, a larger 5.0-kb fragment, encompassing the 3.7-kb region, was active in both fetal and adult definitive hematopoietic cells. This included Ter119⁺ erythroid cells along with fetal liver erythroid and myeloid progenitors. Unlike two other SCL hematopoietic enhancers (+18/19 and −4), +40 enhancer transgenes were inactive in the endothelium. A conserved 400-bp core region, essential for both hematopoietic and midbrain +40 enhancer activity in embryos, relied on two GATA/E-box motifs and was bound in vivo by GATA-1 and SCL in erythroid cells. These results suggest a model in which the SCL +18/19 and/or −4 enhancers initiate SCL expression in early mesodermal derivatives capable of generating blood and endothelium, with subsequent activation of the +40 enhancer via an autoregulatory loop.

We acknowledge and thank the following: Paula Braker, Sarah Pedley, James Harrison, and Denise Weekly for expert generation and care of the transgenic mice; Eric Delabesse for important intellectual and practical contributions to this work, particularly at its inception; Scott Oldham and Beverley Haynes for valuable help with histology; Mike Chapman and Ian Donaldson for help with bioinformatics; Linda Scott for help with hematopoietic colony assays; and Catherine Porcher for the anti-SCL antibody.

This work was supported by The Wellcome Trust.

This work is dedicated to the memory of Isabelle Anne Bouhon (1969 to 2005), a valued friend and colleague.