Differentially active and conserved neural enhancers define two forms of adaptive non-coding evolution in humans
Understanding the molecular processes that underlie human phenotypes is one of the great challenges in modern biology. Comparative genomic studies with our closest living evolutionary relatives, chimpanzees, show a high level of sequence similarity between our species implicating differential gene regulation as an important mechanism contributing to phenotypic divergence. Changes in non-coding regulatory elements are associated with differential traits between species, and non-coding sequence variants are associated with human phenotypic variation in health and disease. Sequences of DNA with putative regulatory function called cis-regulatory elements (CREs) can be identified by computational or experimental methods, and although we can identify these regions computationally, their functions are poorly characterized. In this study we selected human and chimpanzee CREs for functional testing with signs of accelerated evolution in humans or that are active in human brain development. Using a lentiviral-based massively parallel reporter assay (MPRA), we tested the ability of orthologous human and chimpanzee CREs to activate transcription in iPSC-derived neural progenitor cells and neurons. With this assay we identified 179 CREs with differential activity between human and chimpanzee orthologs and 722 CREs with signs of positive selection in humans. Selection and DE CREs differ in level of expression, size, and genomic location. We found a subset of 69 CREs in loci with genetic variants associated with neuropsychiatric diseases, which underscores the consequence of regulatory activity in these loci for proper neural development and function. Unexpectedly, we also found an abundance of CREs with signs of positive selection or differential expression around genes that modulate the Wnt signaling pathway. Together, this analysis of transcriptional activity and selection in CREs uncovers putatively adaptive changes in gene regulation contributing to human specific neural phenotypes.