Harper Richart, Xu Changhong, Di Peter, Chen Yin, Privalsky Martin, Wu Reen
Center for Comparative Respiratory Biology and Medicine, University of California, Davis, CA 95616, United States.
Biochem Biophys Res Commun. 2004 Nov 5;324(1):199-204. doi: 10.1016/j.bbrc.2004.09.037.
Using cDNA microarray analysis, we identified a cDNA clone, DD74, from primary human bronchial epithelial cells, which exhibits increased expression in vitro after treatment with all-trans retinoic acid. This clone corresponded to MAGE D2 mRNA, a gene previously identified to be upregulated in several cancer tissues. Surprisingly, in situ hybridization of lung tissue demonstrated positive hybridization signals with sense, but not antisense, MAGE D2-specific cRNA probes. Examination of several cell lines by Northern blot hybridization confirmed significant expression of two RNA bands. With strand-specific riboprobes, we identified a 2.0kb RNA transcript with the antisense probe as expected and identified a 4.1kb transcript by the sense probe. Further sequence analysis of the 4.1kb transcript revealed at least a 509 nucleotide sequence exactly complementary to the 2.0kb MAGE D2 mRNA sequence. This MAGE D2i sequence contains unique structural features not shared with those of previously described antisense transcripts. Identification of this transcript potentially has important implications for future studies examining MAGE D2 expression patterns in cancer and normal tissues.
通过cDNA微阵列分析，我们从原代人支气管上皮细胞中鉴定出一个cDNA克隆DD74，用全反式维甲酸处理后，它在体外的表达增加。该克隆对应于MAGE D2 mRNA，这是一个先前已鉴定出在几种癌组织中上调的基因。令人惊讶的是，肺组织的原位杂交显示，与正义MAGE D2特异性cRNA探针杂交呈阳性信号，而与反义探针杂交则无阳性信号。通过Northern印迹杂交对几种细胞系进行检测，证实有两条RNA条带表达显著。使用链特异性核糖探针，我们如预期的那样用反义探针鉴定出一个2.0kb的RNA转录本，并用正义探针鉴定出一个4.1kb的转录本。对4.1kb转录本的进一步序列分析显示，至少有一个509个核苷酸的序列与2.0kb MAGE D2 mRNA序列完全互补。这个MAGE D2i序列含有一些独特的结构特征，这些特征是先前描述的反义转录本所没有的。鉴定出这个转录本可能对未来研究MAGE D2在癌组织和正常组织中的表达模式具有重要意义。
