Shaping modern human skull through epigenetic, transcriptional and post-transcriptional regulation of the RUNX2 master bone gene

Abstract

RUNX2 encodes the master bone transcription factor driving skeletal development in vertebrates, and playing a specific role in craniofacial and skull morphogenesis. The anatomically modern human (AMH) features sequence changes in the RUNX2 locus compared with archaic hominins’ species. We aimed to understand how these changes may have contributed to human skull globularization occurred in recent evolution. We compared in silico AMH and archaic hominins’ genomes, and used mesenchymal stromal cells isolated from skull sutures of craniosynostosis patients for in vitro functional assays. We detected 459 and 470 nucleotide changes in noncoding regions of the AMH RUNX2 locus, compared with the Neandertal and Denisovan genomes, respectively. Three nucleotide changes in the proximal promoter were predicted to alter the binding of the zinc finger protein Znf263 and long-distance interactions with other cis-regulatory regions. By surface plasmon resonance, we selected nucleotide substitutions in the 3’UTRs able to affect miRNA binding affinity. Specifically, miR-3150a-3p and miR-6785-5p expression inversely correlated with RUNX2 expression during in vitro osteogenic differentiation. The expression of two long non-coding RNAs, AL096865.1 and RUNX2-AS1, within the same locus, was modulated during in vitro osteogenic differentiation and correlated with the expression of specific RUNX2 isoforms. Our data suggest that RUNX2 may have undergone adaptive phenotypic evolution caused by epigenetic and post-transcriptional regulatory mechanisms, which may explain the delayed suture fusion leading to the present-day globular skull shape.

Mother and Daughter Carrying of the Same Pathogenic Variant in FGFR2 with Discordant Phenotype

“…Each sample was evaluated in triplicate and three independent technical replicates were performed. The expression of the RUNX2 (Runt-related transcription factor 2) gene was analyzed at each time point (24 h, 48 h and 6 days), using semi-quantitative real-time PCR, as already described [ 25 ]. Briefly, qPCR was performed using the GoTaq(R) qPCR Master Mix (Promega) and expression levels were normalized using ACTB (β-actin) as a reference housekeeping gene.…”

Evolutionary mechanisms modulating the mammalian skull development

“…Moreover, changes in the regulation of Runx2 expression during human evolution were linked to skull globularization (i.e. more rounded, less elongated skull) and a delay in cranial suture closure in modern humans [ 128 ]. Runx2 activity levels and the resulting heterochrony in suture closure may account for higher postnatal brain growth rates in early hominids, facilitating encephalization (i.e.…”

Single Nucleotide Polymorphisms in RUNX2 and BMP2 contributes to different vertical facial profile

“…Recent studies exploring SNPs in genes associated with bone physiology have gained attention due to the possible impact on craniofacial growth [ 8 , 10 , 13 , 14 ]. RUNX2 (Runt-related transcription factor 2) gene encodes the master bone transcription factor, crucial for osteoblast differentiation, driving skeletal development and important for craniofacial and dental morphogenesis in vertebrates [ 15 ]. RUNX2 is a downstream target of bone morphogenetic proteins (BMP) family members [ 16 ].…”