Multiple promoters regulate tissue‐specific alternative splicing of the human kallikrein gene, KLK11/hippostasin
Multiple promoters regulate tissue‐specific alternative splicing of the human kallikrein gene, KLK11/hippostasin
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DOI：
10.1111/j.1742-4658.2006.05372.x
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发表时间：
2006-08
期刊：
影响因子：
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通讯作者：
S. Mitsui;Terukazu Nakamura;A. Okui;K. Kominami;H. Uemura;N. Yamaguchi
中科院分区：
文献类型：
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作者：
S. Mitsui;Terukazu Nakamura;A. Okui;K. Kominami;H. Uemura;N. Yamaguchi
KLK家族由位于人类染色体19q13.4上的15个基因组成。KLK11/ hippostasin1是kallikrein家族的成员，在多种组织中表达。从cDNA序列中预测了两种类型的KLK11亚型，即亚型1和亚型2。异构体1已从人海马中分离出来，而异构体2已从前列腺中分离出来。然而，这些异构体的调节和特征是未知的。我们通过确定转录起始位点确定了前三个外显子（1a、1b和1c）。外显子1b包含异构体2的起始密码子，非编码外显子1a和1c贡献了异构体1 mRNA。双荧光素酶启动子分析显示三个启动子区域，对应于每个异构体的第一个外显子。反转录和PCR显示外显子1a在海马、丘脑和非中枢神经系统（CNS）组织中表达，而外显子1b仅在非中枢神经系统组织中检测到。除唾液腺外，在CNS和非CNS组织中均观察到外显子1c。体外诱变表明，1b外显子2亚型的起始密码子是功能性的。同种异构体2在氨基末端有额外的亲水性氨基酸，由神经母细胞瘤细胞系Neuro2a分泌。与绿色荧光蛋白（GFP）融合的异构体1分布在细胞过程中，而异构体2-GFP保留在高尔基体中。我们认为，不仅选择性剪接，而且多种组织特异性启动子的使用也调节了KLK11/hippostasin亚型的表达和细胞内运输。
The human kallikrein (KLK) family consists of 15 genes located on human chromosome 19q13.4. KLK11/hippostasinis a member of the kallikrein family and is expressed in various tissues. Two types of KLK11 isoforms, isoform 1 and isoform 2, have been predicted from cDNA sequences. Isoform 1 has been isolated from human hippocampus, whereas isoform 2 has been isolated from prostate. However, the regulation and characteristics of these isoforms are unknown. We identified the first three exons (1a, 1b, and 1c) by determining their transcription initiation sites. Exon 1b contained the initiation codon of isoform 2, and noncoding exons 1a and 1c contributed to isoform 1 mRNA. The dual luciferase promoter assay revealed three promoter regions, corresponding to the first exon of each isoform. Reverse transcription and PCR showed that exon 1a was expressed in the hippocampus, thalamus, and non‐central nervous system (CNS) tissues, whereas exon 1b was detected only in non‐CNS tissues. Exon 1c was observed in both CNS and non‐CNS tissues, except for salivary glands. In vitro mutagenesis revealed that the initiation codon for isoform 2 in exon 1b was functional. Isoform 2 had additional hydrophilic amino acids at the amino terminal and was secreted from the neuroblastoma cell line Neuro2a. Isoform 1 fused with green fluorescent protein (GFP) was distributed to cellular processes, whereas isoform 2–GFP was retained in the Golgi apparatus. We suggest that not only alternative splicing but also tissue‐specific use of multiple promoters regulate the expression and intracellular trafficking of KLK11/hippostasin isoforms.