Dynamic 3D chromatin architecture contributes to enhancer specificity and limb morphogenesis

The regulatory specificity of enhancers and their interaction with gene promoters is thought to be controlled by their sequence and the binding of transcription factors. By studying Pitx1, a regulator of hindlimbdevelopment, we show that dynamic changes in chromatinconformation can restrict the activity of enhancers. Inconsistent with its hindlimb-restricted expression, Pitx1 is controlled by an enhancer (Pen) that shows activity in forelimbsand hindlimbs. By Capture Hi-C and three-dimensional modeling of the locus, we demonstrate that forelimbsand hindlimbshave fundamentally different chromatinconfigurations, whereby Pen and Pitx1 interact in hindlimbsand are physically separated in forelimbs. Structural variants can convert the inactive into the active conformation, thereby inducing Pitx1 misexpression in forelimbs, causing partial arm-to-leg transformation in mice and humans. Thus, tissue-specific three-dimensional chromatinconformation can contribute to enhancer activity and specificity in vivo and its disturbance can result in gene misexpression and disease.