THE MOUSE B-RAF GENE ENCODES MULTIPLE PROTEIN ISOFORMS WITH TISSUE-SPECIFIC EXPRESSION
THE MOUSE B-RAF GENE ENCODES MULTIPLE PROTEIN ISOFORMS WITH TISSUE-SPECIFIC EXPRESSION
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DOI：
10.1074/jbc.270.40.23381
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发表时间：
1995-10-06
影响因子：
4.8
通讯作者：
CALOTHY, G
中科院分区：
文献类型：
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作者：
BARNIER, JV;PAPIN, C;CALOTHY, G
c-Rmil/B raf原癌基因是编码丝氨酸/苏氨酸蛋白激酶的mil/raf家族的成员，所述丝氨酸/苏氨酸蛋白激酶显示参与从膜到核的信号转导。我们从小鼠脑文库中分离出B-raf cDNA，该cDNA含有先前未鉴定的位于外显子8和9之间的36个碱基对的选择性剪接外显子，因此命名为外显子8b。人和小鼠B raf mRNA还含有先前在禽类c-Rmil基因中描述的120个碱基对的选择性剪接外显子10。位于保守区2（CR2）和催化结构域（CR 3）之间的这两个外显子的独立剪接产生可能编码四种不同蛋白质的mRNA。通过使用针对B-Raf的不同部分产生的特异性血清，我们在成年小鼠组织中鉴定了至少10种蛋白质同种型。一些在69-72 kDa范围内的同种型不被针对外显子1和2编码的肽的抗血清识别，表明存在具有两个不同NH 2末端的B-Raf蛋白。其他亚型，在79-99 kDa的范围内，含有由外显子1和2编码的氨基酸，由一个或两个选择性剪接的外显子编码的氨基酸，并且可能由另一个未鉴定的外显子编码的氨基酸。通过逆转录聚合酶链反应分析B-Raf mRNA表达，并对成年小鼠不同组织中的B-Raf蛋白进行免疫表征，结果显示B-Raf亚型表达具有组织特异性模式。有趣的是，含有外显子10编码的氨基酸的同种型在神经组织中特异性表达。两者合计，这些结果表明，不同的B-Raf蛋白可能参与，在组织特异性的方式，在信号转导途径。
The c-Rmil/B raf proto-oncogene is a member of the mil/raf family encoding serine/threonine protein kinases shown to be involved in signal transduction from the membrane to the nucleus. We isolated from a mouse brain library B-raf cDNAs containing a previously unidentified 36-base pair alternatively spliced exon located between exons 8 and 9 and, therefore, designated exon 8b. Human and mouse B raf mRNAs also contain the 120-base pair alternatively spliced exon 10 previously described in the avian c-Rmil gene. Independent splicing of these two exons, located between the conserved region 2 (CR2) and the catalytic domain (CR3) gives rise to mRNAs potentially encoding four distinct proteins. By using specific sera generated against different portions of B-Raf, we identified at least 10 protein isoforms in adult mouse tissues. Some isoforms, in the range of 69-72 kDa, are not recognized by antisera directed against peptides encoded by exons 1 and 2, indicating the existence of B-Raf proteins with two different NH2 extremities. The other isoforms, in the range of 79-99 kDa, contain the amino acids encoded by exons 1 and 2, by either or both of the alternatively spliced exons, and, possibly, by another unidentified exon. Analysis of B-raf mRNA expression by reverse transcriptase polymerase chain reaction and immunocharacterization of B-Raf proteins in different tissues of the adult mouse showed a tissue-specific pattern of B-Raf isoforms expression. Interestingly, isoforms containing amino acids encoded by exon 10 are specifically expressed in neural tissues. Taken together, these results suggest that distinct B-Raf proteins could be involved, in a tissue-specific manner, in signal transduction pathways.