Cloning of Neuron Specific Protein, MxR

通过筛选小鼠脑内基因文库，克隆了编码小鼠神经内分泌特异性蛋白/网状蛋白(RTN)3的基因。MRTN3/mMxR1的全长为1745nts，含711nts或开放阅读框(ORF)。它有832nls的长3‘非编码区(NCR)，Poly A信号(AATAAA)位于1694。序列分析表明，mRTN3/mMxR1在其C末端与人RTN有很强的同源性。Northern印迹分析表明mRTN3/mMxR1在脑内有表达。它的表达不受胚胎发育的控制。我们在COS-7和HeLa细胞中以融合蛋白的形式表达了ORF。表达的融合蛋白定位于内质网。当唯一的N-末端序列被去除时，截短的融合蛋白仍保留在内质网中，尽管蛋白的强度降低。我们现在正试图确定mRTN3/mMxR1与ER膜的拓扑结构。我们还试图通过合成多肽免疫兔子来制造一种特异的抗体。

We cloned a cDNA encoding mouse neuroendocrine specific protein/reticulon (RTN) 3 by screening mice brain cDNA library. The cDNA of mRTN3/mMxR1 was 1745nts and contains 711nts or open reading frame (ORF). It has 832nls of long 3' noncoding region (NCR), and poly A signal (AATAAA) is located at l694. The sequence analysis indicated that mRTN3/mMxR1 has strong homology with human RTN in its C-terminus. Northern blot analysis indicated that mRTN3/mMxR1 was expressed in brain. Its expression was not controlled embryonic-developmentally. We expressed the ORF as a fusion protein with green fluorescent protein in COS 7 or HeLa cells. The expressed fusion protein was localized in the endoplasmic reticulum. When the unique N-terminal sequence was removed, the truncated fusion protein remained in the ER though the intensity of the protein was reduced. We are now trying to determine the topology of mRTN3/mMxR1 to the ER membrane. We are also trying to make a specific antibodies by immunizing rabbits with synthetic peptides.