Identification of a regulatory domain controlling the Nppa-Nppb gene cluster during heart development and stress

The paralogous genes Nppa and Nppb are organized in an evolutionarily conserved cluster and provide a valuable model for studying co-regulation and regulatory landscape organization during heart development and disease. Here, we analyzed the chromatin conformation, epigenetic status and enhancer potential of sequences of the Nppa-Nppb cluster in vivo. Our data indicate that the regulatory landscape of the cluster is present within a 60-kb domain centered around Nppb. Both promoters and several potential regulatory elements interact with each other in a similar manner in different tissues and developmental stages. The distribution ofH3K27ac and the association of Pol2 across the locus changed during cardiac hypertrophy, revealing their potential involvement in stress-mediated gene regulation. Functional analysis of double-reporter transgenic mice revealed that Nppa and Nppb share developmental, but not stressresponse, enhancers, responsible for their co-regulation.Moreover, the Nppb promoterwas required, but not sufficient, for hypertrophy-induced Nppa expression. In summary, the developmental regulation and stress response of the Nppa-Nppb cluster involve the concerted action of multiple enhancers and epigenetic changes distributed across a structurally rigid regulatory domain.

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