Gao C Y, Chauthaiwale V M, Rampalli A M, Zelenka P S
Laboratory of Molecular and Developmental Biology, National Eye Institute, N.I.H., Bethesda, MD 20932-2730, USA.
Gene. 1996 Oct 17;176(1-2):243-7. doi: 10.1016/0378-1119(96)83274-4.
A rat PCTAIRE-1 cDNA clone was isolated by immunoscreening of a PC12 cDNA library, followed by 5' RACE (rapid amplification of cDNA ends) to determine the 5' end. The rat PCTAIRE-1 cDNA sequence is 96% identical to mouse PCTAIRE-1 and contains an alternatively spliced exon of 131 bp near the 5' end. Although a mouse cDNA containing this exon has been reported, examination of several mouse cell lines provided no evidence for expression of the corresponding mRNA (Okuda et al., 1992). In contrast, reverse transcription and polymerase chain reaction (RT/PCR) across this region using RNA from proliferating, differentiated, and apoptotic PC12 cells demonstrated that alternatively spliced forms of PCTAIRE-1 mRNA with and without this exon are expressed. Both forms of PCTAIRE-1 mRNA are also expressed in vivo in neonatal rat brain, although other tissues examined contained only the form lacking the alternatively spliced exon. In the absence of the alternatively spliced exon PCTAIRE-1 mRNA contains an open reading frame of 1488 bp, corresponding to a 55-kDa protein that is 97% identical to mouse PCTAIRE-1 protein. When the alternatively spliced exon is present, this open reading frame is terminated by a stop codon and a second open reading frame is initiated, predicting a second PCTAIRE-1 protein of 52 kDa. The two predicted PCTAIRE-1 proteins are identical downstream of the splice site, but share no homology at their N-terminal ends.
通过对PC12 cDNA文库进行免疫筛选，分离出大鼠PCTAIRE-1 cDNA克隆，随后进行5' RACE（cDNA末端快速扩增）以确定5' 末端。大鼠PCTAIRE-1 cDNA序列与小鼠PCTAIRE-1的一致性为96%，并且在5' 末端附近含有一个131 bp的可变剪接外显子。尽管已报道了含有该外显子的小鼠cDNA，但对几种小鼠细胞系的检测未发现相应mRNA表达的证据（奥库达等人，1992年）。相反，使用来自增殖、分化和凋亡的PC12细胞的RNA对该区域进行逆转录和聚合酶链反应（RT/PCR）表明，表达了含有和不含有该外显子的PCTAIRE-1 mRNA可变剪接形式。两种形式的PCTAIRE-1 mRNA在新生大鼠脑内也有体内表达，尽管所检测的其他组织仅含有缺乏可变剪接外显子的形式。在没有可变剪接外显子的情况下，PCTAIRE-1 mRNA含有一个1488 bp的开放阅读框，对应于一个55 kDa的蛋白质，该蛋白质与小鼠PCTAIRE-1蛋白质的一致性为97%。当存在可变剪接外显子时，这个开放阅读框由一个终止密码子终止，并启动第二个开放阅读框，预测有一个52 kDa的第二个PCTAIRE-1蛋白质。这两种预测的PCTAIRE-1蛋白质在剪接位点下游是相同的，但在其N末端没有同源性。
