Methyltransferase G9A Regulates Osteogenesis viaTwistGene Repression

Abstract

Here we investigate the role of epigenetic factors in controlling the timing of cranial neural crest cell differentiation. The gene coding for histone H3 lysine 9 methyltransferase G9A was conditionally deleted in neural crest cells with Wnt1-Cre. The majority of homozygous-null animals survived to birth but thereafter failed to thrive. Phenotypic analysis of postnatal animals revealed that the mutants displayed incomplete ossification and 20% shorter jaws as compared to their wild-type littermates. At E13.5, patterns of expression of the osteogenic transcription factor RUNX2 and the mesenchymal transcription factor TWIST are similar in controls and mutants; both overlap in areas of future intramembranous bone formation. At E14.5, the nonosteogenic mesenchyme expressed TWIST, whereas the ossification center had strong RUNX2 and osteopontin expression. In the mutants, TWIST protein was present in the osteogenic mesenchyme, while osteopontin was not expressed until E15.5. In addition, in mutants, small regions of TWIST-positive osteogenic mesenchyme were visible until E15.5. The delay in ossification and reduction in size of the ossification centers were correlated with an earlier decrease in proliferation. We used micromass cultures of the face to investigate the direct effects of G9A inhibition on skeletal differentiation. Addition of a small molecule inhibitor for G9A, BIX-01294, to wild-type cells upregulated Twist genes similar to what was observed in vivo. The inhibitor also caused decreases in several osteogenic markers. Chromatin immunoprecipitation analysis of primary osteogenic mesenchyme from calvaria revealed that Twist1 and Twist2 regulatory regions contain the repressive H3K9me2 marks catalyzed by G9A, which are removed when BIX-01294 is added. Our results establish a role for G9A and H3K9me2 in the regulation of Twist genes and provide novel insights into the significance of epigenetic mechanisms in controlling temporal and tissue-specific gene expression during development.

Human Papillomavirus 16 (HPV16) E2 Repression of TWIST1 Transcription Is a Potential Mediator of HPV16 Cancer Outcomes

“…Rather, E2 binds to the TWIST1 promoter and actively represses transcription from this region. E2 also increases H3K9me2 at the promoter, a hallmark of transcriptionally repressed genes previously implicated in regulation of TWIST1 transcription (53). To our knowledge, this is the first time E2 has been demonstrated to directly bind to a host gene promoter and induce repressive epigenetic markers.…”

“…We next investigated whether the levels of repressive chromatin markers implicated in TWIST1 regulation are changed in the presence of E2. H3K9me2 is a repressive chromatin marker involved in the regulation of TWIST1 expression (53,54), and the levels of this marker on the TWIST1 promoter are increased in N/Tert-11E2 and N/Tert-11HPV16 compared with N/Tert-11Vec ( Fig. 4D; compare lanes 2 and 3, respectively, with lane 1).…”

Cross-talk Between Histone and DNA Methylation Mediates Bone Loss in Hind Limb Unloading

“…But, it has been previously reported that G9a could increase osteogenesis and intramembranous ossification. ( 36 ) The reason for the different effects of G9a on osteogenic activity remains obscure. Depletion of G9a in breast cancer cells could result in both positive and negative effects on steroid hormone‐regulated gene expression.…”

Epidrugs: novel epigenetic regulators that open a new window for targeting osteoblast differentiation

“…They found that these effects were mediated by hyperacetylation of whole chromatin and H3K4 hypermethylation of these genes [ 53 ]. Higashihori et al reported that H3K9 mono- and di-methyltransferase G9a was involved in osteogenesis regulation by twist gene repression [ 54 ]. HDMs have been shown to play an important role in osteo-differentiation of MSCs isolated from patients with oculo-facio-cardio-dental (OFCD) syndrome.…”