Human-derived alleles in SOST and RUNX2 3'UTRs cause differential regulation in a bone cell-line model

The inquiry into the phenotypic features that set apart human species, such as a light, gracile skeleton and a rounded skull characteristic of Homo sapiens, can now benefit from the examination of ancient genomes. These have added a new layer of analysis allowing the identification of genetic differences between species like ours and our closest extinct relatives. Most of these genetic differences are non-coding changes with unknown functional consequences, and dissecting their putative regulatory effect remains challenging. Here we focus on the three prime untranslated regions (3UTR), known to play a critical role in messenger RNA regulation and a plausible locus for divergent regulation between Homo species. We report a set of genes with derived 3UTR changes in either the Homo sapiens or the Neanderthal/Denisovan lineages and experimentally evaluate the impact of 3UTR variants in four genes: E2F6, GLI3, RUNX2 and SOST. We performed a luciferase reporter assay in a bone cell-line model and found a statistically significant difference for the 3UTR variants of SOST (Homo sapiens-derived) and RUNX2 (Neanderthal/Denisovan-derived). The differential expression caused by these variants in our experimental model points to species differences in bone mineral density. Thus, this study adds insights into the functional effects of regulatory variants that emerged in recent human evolution.