RNA secondary structure:: An important cis-element in rat calcitonin/CGRP pre-messenger RNA splicing
RNA secondary structure:: An important cis-element in rat calcitonin/CGRP pre-messenger RNA splicing
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DOI：
10.1021/bi9808058
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发表时间：
1998-11-10
期刊：
影响因子：
2.9
通讯作者：
Roesser, JR
中科院分区：
文献类型：
--
作者：
Coleman, TP;Roesser, JR
降钙素/CGRP基因前mRNA以组织特异性方式选择性拼接，导致甲状腺c细胞中降钙素mRNA和神经元中CGRP mRNA的形成。对含有降钙素特异性外显子4的3'剪接受体的RNA的计算机分析预测，该区域有可能形成一个热力学稳定的茎环。CD光谱和单链特异性和双链特异性RNA酶的溶液相结构探测数据表明，该区域的RNA基本上是双链的。HeLa核提取物中嵌合人β -珠蛋白/降钙素转录物的体外剪接被预测会破坏茎的稳定性的碱基变化所抑制，而代偿性碱基变化导致剪接的水平为野生型的50%。将茎中AG二核苷酸腺苷对面的残基从G改变为U，允许A-U碱基对的形成，在体外消除了这种剪接受体的使用。这些结果表明，在体外降钙素/CGRP基因转录物的外显子4的3'剪接受体上形成了一个热动力学稳定的RNA茎环。这种RNA二级结构作为一种新的顺式元件参与了正确的剪接位点选择。
The calcitonin/CGRP gene pre-mRNA is alternatively spliced in a tissue-specific manner resulting in the formation of calcitonin mRNA in thyroid C-cells and CGRP mRNA in neurons. Computer analysis of the RNA containing the 3' splice acceptor of the calcitonin-specific exon 4 predicts that this region has the potential to form a thermodynamically stable stem-loop. Data from CD spectroscopy and solution phase structure probing with single-strand specific and double-strand specific RNases indicates that RNA in this region is substantially double stranded. In vitro splicing of chimeric human beta-globin/calcitonin transcripts in HeLa nuclear extract was inhibited by base changes predicted to destabilize the stem, while compensatory base changes resulted in splicing at 50% of wild-type levels. Changing the residue opposite the AG dinucleotide adenosine in the stem from G to U, allowing the formation of an A-U basepair, abolished usage of this splice acceptor in vitro. These results indicate that a thermodynamically stable RNA stem-loop forms in vitro at the 3' splice acceptor of exon 4 of the calcitonin/CGRP gene transcript. This RNA secondary structure acts as a novel cis-element involved in proper splice site selection.