3' end formation of human type I and II collagen mRNAs

描述（由申请人提供）：我们对真核mRNA 3'端如何形成有长期的兴趣。基因表达的调节可以通过调节多聚腺苷酸化来完成。我们已经结合了这种兴趣，在多聚腺苷酸化的兴趣，在人类胶原蛋白I型和II型。胶原蛋白在正常软骨发育和骨骼形态发生中起关键作用。我们最近发现并发表了人COL 1A 2多聚腺苷酸化受COL 1A 2 mRNA的3'非翻译区（3' UTR）中存在的顺式作用元件调节，并且这些元件影响3'末端加工效率。我们现在发现的人COL 1A 1和COL 2A 1也含有这些顺式作用元件。基于这些发现，本项目提出研究所有这些I型和II型胶原蛋白前体mRNA的多聚腺苷酸化的调节机制。这些研究将扩大我们的目标，更好地了解胶原蛋白的表达，以及突变和异常基因表达调控如何启动疾病状态。本课题的具体目标是：1）鉴定人胶原基因COL 1A 1、COL 1A 2和COL 2A 1中所有参与3'多聚腺苷酸化位点使用的顺式作用元件; 2）确定与这些顺式作用元件结合的反式作用因子。首先，我们将通过缺失和定点突变来鉴定顺式作用元件，并研究它们在体外和体内影响聚腺苷酸化效率的能力。其次，蛋白质的因素，结合这些顺式作用的信号将进行探索和表征。这些蛋白质因子的确定将为我们提供线索，顺式作用的辅助元件的功能意义。了解这些人胶原基因3'端形成调控中的RNA信号和蛋白因子将有助于我们理解胶原基因表达调控，这反过来又会导致对骨骼缺陷和其他胶原相关疾病的新见解。

DESCRIPTION (provided by applicant): We have a long-standing interest in how eukaryotic mRNA 3' ends are formed. Regulation of gene expression can be accomplished by regulating polyadenylation. We have combined this interest in polyadenylation with an interest in human collagen types I and II. Collagen plays a key role in normal cartilage development and skeletal morphogenesis. We have recently discovered and published that human COL1A2 polyadenylation is regulated by cis-acting elements present in the 3' untranslated region (3' UTR) of the COL1A2 mRNA, and that these elements influence the 3' end processing efficiency. Human COL1A1 and COL2A1 we have now discovered also contain these cis-acting elements. Based upon these findings, this project proposes to investigate the mechanism of regulation of polyadenylation of all these type I and II collagen pre-mRNAs. These investigations will expand our goal of better understanding the expression of collagens, and how mutations and abnormal gene expression regulation can initiate a disease state. Our specific aims for this project are: 1) To identify all the cis-acting elements involved in 3' polyadenylation site usage in the human collagen genes COL1A1, COL1A2, and COL2A1 and 2) To determine what trans-acting factor(s) bind to these cis-acting elements. First, we will identify cis-acting elements by deletion and site-directed mutagenesis, and examine their ability to affect polyadenylation efficiency both in vitro and in vivo. Second, protein factors that bind to these cis-acting signals will be explored and characterized. Determination of these protein factors will provide us with clues as to the functional significance of cis-acting auxiliary elements. Gaining an understanding of the RNA signals and protein factors involved in regulation of 3' end formation of these human collagen genes will help us to comprehend collagen gene expression regulation, which will in turn lead to novel insights into skeletal defects and other collagen-based diseases.