Vachtenheim J, Novotná H
Laboratory of Molecular Biology, University Hospital, 3rd Medical Faculty, Charles University, Prague 8-Bulovka, Czech Republic.
Cell Mol Biol (Noisy-le-grand). 1999 Nov;45(7):1075-82.
Microphthalmia (MITF) gene product, a transcription factor of the basic-helix-loop-helix type, is thought to play a role in the regulation of genes encoding the enzymes necessary for melanogenesis. These include tyrosinase, TRP-1 and TRP-2. Melanocyte-specific isoform of microphthalmia, MITF-M, is expressed in normal and malignant melanocytes. The presence of two other isoforms of microphthalmia, MITF-A and MITF-H, which differ from MITF-M in the amino-terminus, was demonstrated also in some non-melanocytic lineages. Here we have analyzed the presence of all three known isoforms of MITF mRNA in a panel of 17 human melanoma cell lines by a reverse transcriptase-polymerase chain reaction using isoform-specific primers. While, as expected, the predominant form in melanoma cell lines was MITF-M, low amounts of MITF-A mRNA was found in almost all melanomas, as well as in most of 20 tumor cell lines of the non-melanocyte origin (lung and colon carcinomas, osteosarcomas and neuroblastomas). The expression of MITF-H was not detected, with a few exceptions, in the tested cell lines. Pax3 transcription factor was reported earlier to regulate positively the melanocyte-specific promoter of the MITF gene. We found here that the Pax 3 mRNA was expressed in all melanoma cell lines, even in those that had repressed the MITF-M and were amelanotic. This suggests that additional factors, besides Pax3, are required for the MITF expression. The MSG1 (melanocyte-specific gene 1), a gene originally isolated from melanocytes and containing a strong transcription activation domain, was also found expressed in all melanomas and most non-melanocyte tumor cell lines. Together, these data indicate that the MITF-M isoform is the major type of MITF mRNA present in human melanoma cell lines and show that the expression of the isoform MITF-A and the MSG1 is not restricted to malignant melanocytes and occurs in a wide range of tumor cell lines.
小眼畸形（MITF）基因产物是一种碱性螺旋-环-螺旋型转录因子，被认为在调控黑色素生成所需酶的编码基因中发挥作用。这些酶包括酪氨酸酶、TRP-1和TRP-2。小眼畸形的黑色素细胞特异性异构体MITF-M在正常和恶性黑色素细胞中表达。在一些非黑色素细胞谱系中也证实存在小眼畸形的另外两种异构体MITF-A和MITF-H，它们在氨基末端与MITF-M不同。在这里，我们使用异构体特异性引物，通过逆转录聚合酶链反应分析了17个人类黑色素瘤细胞系中所有三种已知MITF mRNA异构体的存在情况。正如预期的那样，黑色素瘤细胞系中的主要形式是MITF-M，但在几乎所有黑色素瘤以及20种非黑色素细胞来源的肿瘤细胞系（肺癌、结肠癌、骨肉瘤和神经母细胞瘤）中的大多数中都发现了少量的MITF-A mRNA。除少数例外，在所测试的细胞系中未检测到MITF-H的表达。先前报道Pax3转录因子可正向调控MITF基因的黑色素细胞特异性启动子。我们在这里发现，Pax 3 mRNA在所有黑色素瘤细胞系中均有表达，即使在那些抑制了MITF-M且无黑色素生成的细胞系中也是如此。这表明除了Pax3之外，MITF表达还需要其他因素。MSG1（黑色素细胞特异性基因1）是一种最初从黑色素细胞中分离出来的基因，含有一个强大的转录激活结构域，也在所有黑色素瘤和大多数非黑色素细胞肿瘤细胞系中表达。总之，这些数据表明MITF-M异构体是人类黑色素瘤细胞系中存在的主要MITF mRNA类型，并表明异构体MITF-A和MSG1的表达不限于恶性黑色素细胞，而是发生在广泛的肿瘤细胞系中。
