Lynx

Human accelerated regions (HARs) are widely interpreted as developmental enhancers, and human-specific substitutions are known to alter their enhancer activity and transcription-factor binding — but systematically only in the brain. Whether HARs active outside the brain carry substitutions that change binding at experimentally bound transcription-factor sites, distinct from the GC-biased gene conversion (gBGC) that inflates substitution counts at these GC-rich elements, has not been examined. From a single-cell chromatin-accessibility matrix (312 Zoonomia HARs × 16 tissues) we identified 45 dominantly accessible outside the brain and screened each for human-specific substitutions inside experimentally bound (UniBind ChIP-seq) transcription-factor motifs, re-scoring affinity against the reconstructed primate ancestor and testing each change against a per-locus gBGC null. 19 carry a substitution inside a bound motif, and 8 show an affinity gain exceeding the neutral gBGC expectation (P < 0.05). The strongest, ZOOHAR.189, creates a CTCF motif match at an experimentally bound site near MYLK2 (Δ +8.0 bits); others gain bound TP63, GATA3 and PRDM1 sites in tissue-matched contexts. These position-weight-matrix predictions remain hypotheses pending allele-specific assays, but nominate specific loci and tissues at which human substitutions may have rewired regulatory binding.