Krox20 hindbrain regulation involves multiple modes of cooperation between cis-acting elements [ATAC-seq]. Mus musculus

Many genes harbour multiple transcriptional enhancers that act concomitantly to achieve robust and precise spatial-temporal expression. In vertebrates, however, the mechanisms underlying cooperation between cis-acting elements are poorly documented. The mouse gene Krox20 encodes a transcription factor required for the specification of two segments (rhombomeres) of the developing hindbrain. In rhombomere 3, Krox20 is subject to positive feedback, governed by enhancer A, which is directly bound by the KROX20 protein, whereas another element, C, distant from 70 kb, was supposed to be only required for initiation of expression. Here, using both enhancer knock-outs and investigations of chromatin organisation, we show that element C possesses a dual activity: besides its classical enhancer function, it is also permanently required in cis to potentiate element A autoregulatory activity, by increasing its chromatin accessibility. This work uncovers a novel, asymmetrical, long-range mode of cooperation between cis-acting elements that might be essential to avoid promiscuous activation of positive autoregulatory elements. Overall design: Chromatin accessibility assessed by ATAC-seq from mouse embryonic samples at different time points and anatomical positions.