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CHD9 upregulates RUNX2 and has a potential role in skeletal evolution.
BMC Molecular and Cell Biology ( IF 2.4 ) Pub Date : 2020-04-15 , DOI: 10.1186/s12860-020-00270-5
Axel H Newton 1 , Andrew J Pask 1
Affiliation  

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.

中文翻译:

CHD9上调RUNX2并在骨骼发育中具有潜在作用。

背景技术基因调控的变化被广泛认为是适应性表型进化的重要驱动力。但是,仍然缺乏了解支持这种变化的特定分子机制。染色质状态通过影响编码基因座对转录机制的可及性,在基因调控中起着至关重要的作用。因此,染色质重塑剂功能的改变是驱动与表型适应性相关的基因表达改变的强烈候选者。在这里,我们确定了染色质重塑剂CHD9中的氨基酸同质性,它们在灭绝的有袋甲状腺蛋氨酸和以太狼之间共享,显示出明显的颅骨融合。尽管CHD9在该过程中的作用仍知之甚少,但它参与了成骨过程。我们检查CHD9是否能够调节成骨靶基因的表达,并检查CHD9 DNA结合域中关键取代的功能。结果我们检查了CHD9是否能够上调HEK293T细胞中的成骨靶基因RUNX2,骨钙蛋白(OC)和ALP。我们发现,CHD9的过表达上调了成骨细胞命运的主要调节者RUNX2,但没有下游基因OC或ALP,这支持了CHD9调节成骨祖细胞而不是终末成骨细胞的观点。我们还发现,CHD9 DNA结合结构域中的进化取代不会改变蛋白质的二级结构,但是能够驱动RUNX2激活的增加,但微不足道。最后,CHD9无法激活游离型RUNX2启动子-报告子构建体,提示CHD9的功能需要完整的染色质补体。结论我们通过新观察到的CHD9上调RUNX2表达的能力为CHD9在成骨分化中的作用提供了新的证据。尽管我们无法确定HEK293T细胞中进化替代的重要功能后果,但我们的研究为蛋白质同质体的功能研究及其在发育过程中的作用提供了重要的步骤。编码基因中的突变可能是驱动基因表达适应性变化的机制,其验证对于确定进化同源性的功能后果至关重要。结论我们通过新观察到的CHD9上调RUNX2表达的能力为CHD9在成骨分化中的作用提供了新的证据。尽管我们无法确定HEK293T细胞中进化替代的重要功能后果,但我们的研究为蛋白质同质体的功能研究及其在发育过程中的作用提供了重要的步骤。编码基因中的突变可能是驱动基因表达适应性变化的机制,其验证对于确定进化同源性的功能后果至关重要。结论我们通过新观察到的CHD9上调RUNX2表达的能力为CHD9在成骨分化中的作用提供了新的证据。尽管我们无法确定HEK293T细胞中进化替代的重要功能后果,但我们的研究为蛋白质同质体的功能研究及其在发育过程中的作用提供了重要的步骤。编码基因中的突变可能是驱动基因表达适应性变化的机制,它们的验证对于确定进化同源性的功能后果至关重要。我们的研究为蛋白质同源性的功能研究及其在发育过程中的作用提供了重要的步骤。编码基因中的突变可能是驱动基因表达适应性变化的机制,其验证对于确定进化同源性的功能后果至关重要。我们的研究为蛋白质同源性的功能研究及其在发育过程中的作用提供了重要的步骤。编码基因中的突变可能是驱动基因表达适应性变化的机制,它们的验证对于确定进化同源性的功能后果至关重要。
更新日期:2020-04-22
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