Site pleiotropy of a stickleback Bmp6 enhancer

Development and regeneration are orchestrated by gene regulatory networks that operate in part through tran-scriptional enhancers. Although many enhancers are pleiotropic and are active in multiple tissues, little is known about whether enhancer pleiotropy is due to 1) site pleiotropy, in which individual transcription factor binding sites (TFBS) are required for activity in multiple tissues, or 2) multiple distinct sites that regulate expression in different tissues. Here, we investigated the pleiotropy of an intronic enhancer of the stickleback Bone morpho-genetic protein 6 (Bmp6) gene. This enhancer was previously shown to regulate evolved changes in tooth number and tooth regeneration, and is highly pleiotropic, with robust activity in both fins and teeth throughout em-bryonic, larval, and adult life, and in the heart and kidney in adult fish. We tested the hypothesis that the plei-otropy of this enhancer is due to site pleiotropy of an evolutionarily conserved predicted Foxc1 TFBS. Transgenic analysis and site-directed mutagenesis experiments both deleting and scrambling this predicted Foxc1 TFBS revealed that the binding site is required for enhancer activity in both teeth and fins throughout embryonic, larval, and adult development, and in the heart and kidney in adult fish. Collectively these data support a model where the pleiotropy of this Bmp6 enhancer is due to site pleiotropy and this putative binding site is required for enhancer activity in multiple anatomical sites from the embryo to the adult.

Automated summary

Development and regeneration are controlled by gene regulatory networks, including transcriptional enhancers. This study focuses on the pleiotropy of an intronic enhancer of the Bmp6 gene and shows that the enhancer's pleiotropy is due to the site pleiotropy of a predicted Foxc1 TFBS.