Promoter-enhancer looping and shadow enhancers of the mouse αA-crystallin locus

Gene regulation by enhancers is important for precise temporal and spatial gene expression. Enhancers can drive gene expression regardless of their location, orientation or distance from the promoter. Changes in chromatin conformation and chromatin looping occur to bring the promoter and enhancers into close proximity. αA-crystallin ranks among one of the most abundantly expressed genes and proteins in the mammalian lens. The αA-crystallin locus is characterized by a 16 kb chromatin domain marked by two distal enhancers, 5' DCR1 and 3' DCR3. Here we used chromatin conformation capture (3C) analysis and transgenic approaches to analyze temporal control of the mouse αA-crystallin gene. We find that DCR1 is necessary, but not sufficient alone to drive expression at E10.5 in the mouse lens pit. Chromatin looping revealed interaction between the promoter and the region 3' to DCR1, identifying a novel enhancer region in the αA-crystallin locus. We determined that this novel enhancer region, DCR1S, recapitulates the temporal control by DCR1. Acting as shadow enhancers, DCR1 and DCR1S are able to control expression in the lens vesicle at E11.5. It remains to be elucidated however, which region of the αA-crystallin locus is responsible for expression in the lens pit at E10.5.