Position and sequence requirements for poly(A) length regulation by the poly(A) limiting element.
Position and sequence requirements for poly(A) length regulation by the poly(A) limiting element.
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Poly(A) 限制元件对 Poly(A) 长度调节的位置和序列要求。
DOI：
10.1017/s1355838201010329
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Schoenberg,DR
中科院分区：
文献类型：
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作者：
Gupta,JD;Gu,H;Schoenberg,DR
Poly(A)-限制元件(PLE)是一种顺式作用序列，其作用是将Pre-mRNAs上的Poly(A)尾部长度限制在20个核苷酸以内。功能性PLE存在于许多基因中，强调了这种控制机制的普遍性。本研究试图进一步确定Poly(A)长度调控的位置要求和组成PLE的核心序列。增加PLE与上游3‘剪接点之间或PLE与下游AAUAAA之间的距离对Poly(A)长度控制没有影响。然而，将PLE从末端外显子转移到上游外显子或内含子，消除了Poly(A)长度控制。使用一组构建物进一步评估Poly(A)长度控制，其中PLE保持在末端外显子，但上游内含子被删除、修饰或替换为多嘧啶区域。Poly(A)长度控制在所有情况下都保持不变，这表明关键特征是在末端外显子中存在PLE。一系列突变证明了该元素富含5‘嘧啶部分的重要性。最后，UV交联实验在Hela核提取液中发现了一种∼62-kDa蛋白质，它与野生型23-NT PLE RNA寡核苷酸结合，但不与该元素的突变的非功能形式结合。
The poly(A)-limiting element (PLE) is a cis-acting sequence that acts to limit poly(A) tail length on pre-mRNA to <20 nt. Functional PLEs are present in a number of genes, underscoring the generality of this control mechanism. The current study sought to define further the position requirements for poly(A) length regulation and the core sequence that comprises a PLE. Increasing the spacing between the PLE and the upstream 3′ splice site or between the PLE and the downstream AAUAAA had no effect on poly(A) length control. However, moving the PLE from the terminal exon to either an upstream exon or intron eliminated poly(A) length control. Poly(A) length control was further evaluated using a battery of constructs in which the PLE was maintained in the terminal exon, but where upstream introns were either deleted, modified, or replaced with a polypyrimidine tract. Poly(A) length control was retained in all cases, indicating that the key feature is the presence of the PLE in the terminal exon. A battery of mutations demonstrated the importance of the 5′ pyrimidine-rich portion of the element. Finally, UV crosslinking experiments identified an ∼62-kDa protein in Hela nuclear extract that binds to a wild-type 23-nt PLE RNA oligonucleotides but not to a mutated nonfunctional form of the element.