Interrogating the Regulatory Function of HAQERs during Human Cortical Development

Sequence divergence within gene regulatory elements has been proposed to play an important role in the evolution of human-specific traits, including cortical expansion. However, the mutational processes that efficiently modify gene regulatory elements and the target genes upon which they act are poorly understood. We investigated the regulatory function and origins of the fastest evolved regions in the human genome, termed Human Ancestor Quickly Evolved Regions (HAQERs), in their native genomic context during human cerebral cortex development.

We identified 50 HAQERs with accessible chromatin in developing human cortex, largely arising from previously unconstrained ancestral sequences. To test the necessity of these HAQERs for gene regulation, we established an all-in-one CRISPRi lentiviral vector and linked 26 HAQERs to nearby target genes across cell types and Wnt pathway activation contexts. Rapid gains of CpGs distinguished HAQERs active during cortical development and displaying human-specific epigenomic marks. As a high density of CpG sites can drive formation of permissive chromatin, we identified 107 HAQERs with at least 17 human-specific CpG gains per kb, termed HAQER CpG Beacons. These HAQERs emerged via contributions from GC-biased gene conversion (gBGC) with evidence for selection preferentially fixing CpG sites. Notably, the CHL1 and DPP10 loci, both implicated in human neurological disorders, each harbored two gene-linked HAQER CpG Beacons.

Our findings reveal HAQER target genes and support a model where gBGC and natural selection jointly drive regulatory-altering CpG variants to fixation, forging regulatory innovations in the human cortex.