Comparative Genomics Identifies the Mouse <em>Bmp3</em> Promoter and an Upstream Evolutionary Conserved Region (ECR) in Mammals

The Bone Morphogenetic Protein (BMP) pathway is a multi-member signaling cascade whose basic components are found in all animals. One member, BMP3, which arose more recently in evolution and is found only in deuterostomes, serves a unique role as an antagonist to both the canonical BMP and Activin pathways. However, the mechanisms that control BMP3 expression, and the cis-regulatory regions mediating this regulation, remain poorly defined. With this in mind, we sought to identify the Bmp3 promoter in mouse (M. musculus) through functional and comparative genomic analyses. We found that the minimal promoter required for expression in resides within 0.8 kb upstream of Bmp3 in a region that is highly conserved with rat (R. norvegicus). We also found that an upstream region abutting the minimal promoter acts as a repressor of the minimal promoter in HEK293T cells and osteoblasts. Strikingly, a portion of this region is conserved among all available eutherian mammal genomes (47/47), but not in any non-eutherian animal (0/136). We also identified multiple conserved transcription factor binding sites in the Bmp3 upstream ECR, suggesting that this region may preserve common cis-regulatory elements that govern Bmp3 expression across eutherian mammals. Since dysregulation of BMP signaling appears to play a role in human health and disease, our findings may have application in the development of novel therapeutics aimed at modulating BMP signaling in humans.骨形态发生蛋白（Bone Morphogenetic Protein, BMP）通路是一类多成员信号级联反应，其核心组分广泛存在于所有动物类群中。其中一个成员BMP3在进化过程中出现较晚，仅见于后口动物，可作为经典BMP通路与激活素通路的双重拮抗因子，发挥独特的生物学功能。然而，调控BMP3表达的分子机制，以及介导该调控的顺式调控区域（cis-regulatory regions），目前仍未得到充分阐明。鉴于此，我们旨在通过功能基因组学与比较基因组学分析，鉴定小家鼠（Mus musculus）的Bmp3基因启动子。研究发现，介导Bmp3基因表达所需的最小启动子（minimal promoter）位于Bmp3基因上游0.8 kb的区域内，该区域与褐家鼠（Rattus norvegicus）的对应序列具有高度保守性。此外，我们还发现紧邻最小启动子的上游区域可在HEK293T细胞与成骨细胞中对最小启动子发挥阻遏作用。值得注意的是，该区域的一段序列在所有已公开的真兽类哺乳动物基因组中均呈现保守性（47/47），但在所有非真兽类动物中均未发现该保守序列（0/136）。我们还在Bmp3基因上游的进化保守区域（ECR）中鉴定出多个保守的转录因子结合位点（transcription factor binding sites），提示该区域可能保留了调控真兽类哺乳动物Bmp3表达的共同顺式调控元件。鉴于BMP信号通路的失调与人类健康及疾病发生发展密切相关，本研究结果可为开发靶向调控人类BMP信号通路的新型治疗策略提供潜在应用价值。