Regulation of SMAD4 and BMPR1A Expression in Juvenile Polyposis

幼年性息肉病（JP）是一种常染色体显性遗传综合征，易发生结肠、直肠和胃的错构瘤性息肉。受影响的患者有大约50%的风险发展为胃肠道癌。我们实验室的工作已经确定了2个导致JP的基因，这两个基因都是转化生长因子β（TGF-β）超家族的成员。这些基因之一SMAD 4是通过TGF-β、骨形态发生蛋白（BMP）和激活素途径进行信号传导的常见细胞内介质。另一种是BMPR 1A，是一种细胞表面受体，将BMP信号从细胞膜转导到细胞质中。在大约20%的JP病例的生殖系中分别发现了每个基因编码序列的突变。其他60%的JP病例的原因尚不清楚，可能包括其他未发现的易患JP的基因，测序无法检测到的已知基因的较大缺失，以及这些基因非编码区的变化，从而改变基因表达。过去几年我们JP研究的重点是通过对大型JP家族进行基于连锁的基因组筛选来发现新的JP基因，并对TGF-β超级家族中的其他基因进行直接测序，以了解大量JP家族中的突变。我们还探讨了外显子缺失的患病率，占JP病例的4%。最近的一项挑衅性发现来自基因组筛选和删除研究。在一个没有SMAD 4或BMPR 1A编码突变的大JP家族中，发现第三个JP基因不是JP的原因，而是BMPR 1A的推定启动子区和第一个非编码外显子的缺失通过种系遗传。这使得我们将研究重点放在研究已知的JP基因是如何调节并导致BMP信号转导改变的。另一名JP患者被发现缺失SMAD 4的推定启动子和前2个非编码外显子，进一步支持这种方法。因此，在这项研究中，我们建议研究影响JP基因和蛋白质表达的因素，这是一种基本上未探索的和潜在的非常重要的机制，是常染色体显性遗传癌症综合征的遗传基础。我们的具体目标是：1）表征SMAD 4和BMPR 1A的启动子区和非编码外显子，并评估JP患者的生殖系变化; 2）确定JP基因编码区和非编码区突变对RNA和蛋白质表达的影响。

Juvenile Polyposis (JP) is an autosomal dominant syndrome predisposing to the development of hamartomatous polyps of the colon, rectum, and stomach. Affected patients have an approximately 50% risk of developing gastrointestinal cancer. Work in our laboratory has identified 2 genes that cause JP, both of which are members of the transforming growth factor beta (TGF-β) super family. One of these genes, SMAD4, is the common intracellular mediator of signaling through the TGF-β, bone morphogenetic protein (BMP), and activin pathways. The other, BMPR1A, is a cell surface receptor which transduces BMP signals from the cell membrane into the cytoplasm. Mutations in the coding sequence of each gene have been found in the germline of approximately 20% of JP cases, respectively. The cause for the other 60% of JP cases is unknown, and could include other undiscovered genes predisposing to JP, larger deletions of the known genes not detectable by sequencing, and changes in non-coding regions of these genes, thereby altering gene expression. The focus of our JP studies over the past several years has been to discover new JP genes through a linkage-based genome screen of a large JP kindred, and direct sequencing of other genes in the TGF-β super family for mutations in a large number of JP families. We have also explored the prevalence of exonic deletions, which accounted for another 4% of JP cases. A recent provocative finding came from this genome screen and deletion studies. In a large JP family without coding mutations of SMAD4 or BMPR1A, it was discovered that a third JP gene was not the cause of JP, but rather, a deletion of the putative promoter region and first non-coding exon of BMPR1A was inherited through the germline. This has led us to focus our studies upon investigating how the known JP genes are regulated and lead to altered BMP signaling. Another JP patient has been found with a deletion of the putative promoter and first 2 non-coding exons of SMAD4, lending further support to this approach. Therefore, in this grant we propose to examine factors which affect the expression of JP genes and proteins, a largely unexplored and potentially very important mechanism underlying the genetic basis of autosomal dominant cancer syndromes. Our specific aims are: 1) to characterize the promoter region and non-coding exons of SMAD4 and BMPR1A and evaluate JP patients for germline changes; and 2) to determine the influence of mutations in coding and non-coding regions of JP genes on RNA and protein expression.