Study of cell type specific gene expression of runt related murine genes

PEBP2Alpha亚基是一个新的转录因子家族的成员，其结构域负责其与DNA的结合和与β亚基活性的异构化。目前已鉴定出三种小鼠PEBP2pha基因，即αA、αB和αC。AlphaA最初是作为一种多瘤病毒增强子结合蛋白被分离出来的，参与T细胞特异性基因表达或早期发育。Alphab是在t(8，21)断裂点发现的人类AML1的小鼠同源物。我们鉴定了唯一编码β亚基的基因，并且在我们测试的细胞系中普遍存在其mRNA的表达。AlphaA mRNAs的表达提示细胞类型的特异性，并提示改变其作为转录因子的功能。在本研究中，我们分析了alphaA亚种(alphaA1、alphaA2、alphaA3)mRNAs的表达模式，以表征mRNA表达的细胞类型特异性机制。初步的Northern印迹分析表明，AlphaA3mRNA是类似于AlphaA2的次要物种。为了证实它们的表达，RT-PCR发现，在Ha-ras转化的NIH3T3细胞和小鼠T细胞系EL4中都表达了类似于AlphaA1的AlphaA3mRNA。此外，在小鼠胚胎癌细胞系P19中几乎没有检测到αA3和αA1的表达。维甲酸处理后，P19细胞中αA3mRNA的表达显著增加。另一方面，在我们测试的所有细胞系和β亚基mRNAs中都有alphaA2基因的表达。这些研究有力地表明，单个基因通过选择性剪接产生的各种异构体对每个mRNAs的表达有不同的调控。这种表达的特异性及其转录调控机制的生物学意义值得进一步探讨。

PEBP2alpha subunit is a member of a novel transcription factor family with the Runt-Domain which is responsible for its DNA binding and hetrodimerizing with beta subunit activities. Three murine PEBP2alpha genes, alphaA, alphaB and alphaC, have been identified. alphaA is originally isolated as a Polyoma virus enhancer binding protein and involved the T cell specific gene expression or early development. alphaB is the murine homologue of human AML1 which was found at the breakpoint of the t(8,21). The only gene encoding beta subunit was identified and the expression of its mRNA is ubiquitous among the cell lines we tested. The expressions of alphaA mRNAs indicate the cell type specific manner and suggest to vary its function as a transcription factor. In this study, we analyzed the expression pattern of alphaA subspecies (alphaA1 , alphaA2, alphaA3) mRNAs to characterize the machinery of cell type specificity of mRNA expression. A primary Northern blot analysis suggested that alphaA3 mRNA was minor species like alphaA2 mRNA.To confirm their expression, RT-PCR was performed and was found that the alphaA3 mRNA, like alphaA1, was expressed both in Ha-ras transformed NIH3T3 cells and murine T cell line, EL4. Moreover, little expression of alphaA3, like alphaA1, was detected in murine embryonic carcinoma cell line, P19. After the treatment of retinoic acid, however, significant induction of alphaA3 mRNA was obsered in the P19 cells. On the other hand, alphaA2 mRNA was expressed in all cell lines we tested as well as beta subunit mRNAs. These study strongly suggested that the various isoforms derived from a single gene by alternative splicing were differently regulated the expression of each mRNA.It is a valuable question we should discuss further that the biological significance of this expression specificity and the mechanism of their transcription regulation.