CHD9 upregulates RUNX2 and has a potential role in skeletal evolution

Abstract

Background: Changes in gene regulation are widely recognized as an important driver of adaptive phenotypic evolution. However, the specific molecular mechanisms that underpin such changes are still poorly understood. Chromatin state plays an essential role in gene regulation, by influencing the accessibility of coding loci to the transcriptional machinery. Changes in the function of chromatin remodellers are therefore strong candidates to drive changes in gene expression associated with phenotypic adaptation. Here, we identify amino acid homoplasies in the chromatin remodeller CHD9, shared between the extinct marsupial thylacine and eutherian wolf which show remarkable skull convergence. CHD9 is involved in osteogenesis, though its role in the process is still poorly understood. We examine whether CHD9 is able to regulate the expression of osteogenic target genes and examine the function of a key substitution in the CHD9 DNA binding domain.Results: We examined whether CHD9 was able to upregulate its osteogenic target genes, RUNX2, Osteocalcin (OC) and ALP in HEK293T cells. We found that overexpression of CHD9 upregulated RUNX2, the master regulator of osteoblast cell fate, but not the downstream genes OC or ALP, supporting the idea that CHD9 regulates osteogenic progenitors rather than terminal osteoblasts. We also found that the evolutionary substitution in the CHD9 DNA binding domain does not alter protein secondary structure, but was able to drive a small but insignificant increase in RUNX2 activation. Finally, CHD9 was unable to activate an episomal RUNX2 promoter-reporter construct, suggesting that CHD9 requires the full chromatin complement for its function. Conclusions: We provide new evidence to the role of CHD9 in osteogenic differentiation through its newly observed ability to upregulate the expression of RUNX2. Though we were unable to identify significant functional consequences of the evolutionary substitution in HEK293T cells, our study provides important steps forward in the functional investigation of protein homoplasy and its role in developmental processes. Mutations in coding genes may be a mechanism for driving adaptive changes in gene expression, and their validation is essential towards determining the functional consequences of evolutionary homoplasy.

Pterostilbene attenuates the proliferation and differentiation of TNF‑α‑treated human periodontal ligament stem cells

“…A previous study also revealed that PTE inhibited inflammation and apoptosis in mouse models of pulmonary fibrosis (37). ALP has been considered to be an early osteogenic marker, whilst RUNX2 regulates the differentiation of pluripotent stem cells into osteoblasts (38).…”

The Genomes of Two Billfishes Provide Insights into the Evolution of Endothermy in Teleosts

“…The bill of billfishes results from the specialization of the upper jaw bone. Previous studies have found that the gene runx2 was a major driver of craniofacial evolution in mammals, while chd9 was found to be a potential regulator for runx2 ( Newton and Pask 2020 ). Interestingly, among the two CNEs with billfish-specific deletions, one (UCE-1582) is located 172 kb upstream of chd9 and they are also in the same TAD ( fig.…”

Sentinels of chromatin: chromodomain helicase DNA-binding proteins in development and disease

“…For instance, some, such as CHD1, are required for RNAPII transcription ( Krogan et al 2002 ; Simic 2003 ; Srinivasan et al 2005 ; Warner et al 2007 ; Skene et al 2014 ; Dieuleveult et al 2016 ; Baumgart et al 2017 ) and play a role in transcription initiation ( Lin et al 2011 ) and in RNAPII-directed turnover of promoter-proximal nucleosomes, promoter escape, and subsequent elongation and splicing ( Sims et al 2007 ; Skene et al 2014 ). Similarly, mounting evidence shows that mammalian CHD2 ( Rom et al 2019 ), CHD6 ( Lutz et al 2006 ), CHD8 ( Rodríguez-Paredes et al 2009 ; Ceballos-Chávez et al 2015 ), and CHD9 ( Shur et al 2006a ; Lee and Stallcup 2017 ; Alendar et al 2020 ; Newton and Pask 2020 ; Yoo et al 2020 ) potentiate transcription of distinct target genes in diverse cell types. CHD enzymes also contribute to transcriptional repression, best exemplified by the concerted action of chromatin remodeling and histone deacetylase catalytic subunits of the NuRD complex, which generally leads to the generation of hypoacetylated, densely packed chromatin ( Lai and Wade 2011 ) and facilitates PRC2-mediated silencing of target genes in ESCs ( Reynolds et al 2012 ).…”