Transcriptional Enhancers in the FOXP2 Locus Underwent Accelerated Evolution in the Human Lineage

Abstract

Unique human features such as complex language are the result of molecular evolutionary changes that modified developmental programs of our brain. The human-specific evolution of the forkhead box P2 (FOXP2) gene coding region has been linked to the emergence of speech and language in the human kind. However, little is known about how the expression of FOXP2 is regulated and if its regulatory machinery evolved in a lineage-specific manner in humans. In order to identify FOXP2 regulatory regions containing human-specific changes we used databases of human accelerated non-coding sequences or HARs. We found that the topologically associating domain (TAD) determined using developing human cerebral cortex containing the FOXP2 locus includes two clusters of 12 HARs, placing the locus occupied by FOXP2 among the top regions showing fast acceleration rates in non-coding regions in the human genome. Using in vivo enhancer assays in zebrafish, we found that at least five FOXP2-HARs behave as transcriptional enhancers throughout different developmental stages. In addition, we found that at least two FOXP2-HARs direct the expression of the reporter gene EGFP to foxP2 expressing regions and cells. Moreover, we uncovered two FOXP2-HARs showing reporter expression gain of function in the nervous system when compared with the chimpanzee ortholog sequences. Our results indicate that regulatory sequences in the FOXP2 locus underwent a human-specific evolutionary process suggesting that the transcriptional machinery controlling this gene could have also evolved differentially in the human lineage.