Structure, expression, and some regulatory mechanisms of the rat preprotachykinin gene encoding substance P, neurokinin A, neuropeptide K, and neuropeptide gamma
Structure, expression, and some regulatory mechanisms of the rat preprotachykinin gene encoding substance P, neurokinin A, neuropeptide K, and neuropeptide gamma
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DOI：
10.1523/jneurosci.10-07-02203.1990
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发表时间：
1990-07
期刊：
影响因子：
--
通讯作者：
M. Carter;J. Krause
中科院分区：
文献类型：
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作者：
M. Carter;J. Krause
从λ Charon 4A基因组文库中分离出编码神经肽P物质（SP）、神经激肽A（NKA）、神经肽K（NPK）和神经肽γ的大鼠前速激肽原（PPT）基因。两个重叠克隆包含了β-PPT中存在的所有外显子，分别包括约7和9 kb的5′和3′侧翼序列。通过引物延伸和核酸酶保护实验确定了1个主要和2个次要转录起始位点的存在。对大鼠和牛之间的核苷酸序列同源性的分析显示，在整个编码区和5′侧翼序列内存在高度保守的区域。引物延伸和核酸酶保护实验表明，初级转录本差异剪接主要为γ-和β-PPT mRNA在成年大鼠中的基因表达的所有组织检查。β-PPT mRNA包含所有外显子，而γ-PPT mRNA缺少外显子4，其编码NPK的部分N-末端。缺少外显子6（编码NKA和加工位点的序列）的α-PPT mRNA占总PPT mRNA的约1%。提出了一个RNA二级结构模型来解释RNA剪接过程中这些特定的外显子排除事件。这些结果进行了讨论方面的机制调节SP基因的表达和差异RNA剪接在大鼠的功能意义。
The rat preprotachykinin (PPT) gene encoding the neuropeptides substance P (SP), neurokinin A (NKA), neuropeptide K (NPK), and neuropeptide gamma was isolated from a lambda Charon 4A genomic library. Two overlapping clones contained all of the exons present in beta-PPT, including some 7 and 9 kb 5′ and 3′ flanking sequence, respectively. The presence of 1 major and 2 minor transcription initiation sites was determined from primer extension and nuclease protection experiments. Analysis of the nucleotide sequence homology between the rat and bovine revealed the presence of highly conserved regions throughout the entire coding region and within the 5′ flanking sequences. Primer extension and nuclease protection experiments demonstrated that the primary transcript is differentially spliced primarily into gamma- and beta-PPT mRNA in all tissues examined in the adult rat where the gene is expressed. beta-PPT mRNA contains all of the exons, whereas gamma-PPT mRNA lacks exon 4, which encodes part of the N-terminus of NPK. The alpha-PPT mRNA, which lacks exon 6 (the sequence encoding NKA and processing sites), comprises about 1% of the total PPT mRNA. An RNA secondary structure model is proposed to account for these specific exon exclusion events in the RNA splicing process. These results are discussed with regard to the mechanisms regulating SP gene expression and the functional significance of differential RNA splicing in the rat.