Characterization of the isoforms of MOVO zinc finger protein, a mouse homologue of Drosophila Ovo, as transcription factors.
Characterization of the isoforms of MOVO zinc finger protein, a mouse homologue of Drosophila Ovo, as transcription factors.
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DOI：
10.1016/j.gene.2004.03.013
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发表时间：
2004-07
期刊：
影响因子：
3.5
通讯作者：
S. Unezaki;M. Nishizawa;E. Okuda‐Ashitaka;Y. Masu;M. Mukai;Satoru Kobayashi;K. Sawamoto;H. Okano;S. Ito
中科院分区：
文献类型：
--
作者：
S. Unezaki;M. Nishizawa;E. Okuda‐Ashitaka;Y. Masu;M. Mukai;Satoru Kobayashi;K. Sawamoto;H. Okano;S. Ito
我们之前描述了一种新的锌指蛋白MoVo的两种异构体(MoVo-A和-B)，MoVo是果蝇Ovo蛋白的小鼠同源物。在这里，我们克隆了编码第三异构体MoVo-C的cDNA，它有一个反式激活结构域和一个锌指结构域，但缺乏一个存在于MoVo-A中的N末端潜在的阻遏结构域。3种异构体mRNAs在小鼠睾丸中高表达，在卵巢中低水平表达。对分离的MoVo基因和mRNAs的结构分析表明，三种不同的MoVo转录本被差异处理，产生三种异构体。主要信使核糖核酸编码MoVo-B的锌指结构域，次要信使核糖核酸编码MoVo-A(潜在抑制因子)和MoVo-C(潜在激活因子)。为了将MoVo分配给一个转录因子，我们表征了DNA结合和反式激活特性。随机寡核苷酸选择、凝胶迁移率改变分析和足迹分析表明，MoVo与5‘-G(G/C/T)GGGGG-3’序列结合。这些基序存在于MoVo和其他睾丸特异基因的5‘侧翼区。在小鼠睾丸中检测到与该基序结合的核蛋白，在精母细胞中MoVo mRNA的表达受到限制。荧光素酶活性检测表明，MoVo-C激活了MoVo启动子，MoVo-A抑制了MoVo启动子，而MoVo-B无此作用。MoVo启动子中MoVo结合基序的突变降低了荧光素酶的活性。所有异构体对不含MoVo结合基序的SV40启动子均无作用。MoVo-A部分挽救了果蝇卵突变体的卵子发生。这些结果表明，MoVo亚型是调控睾丸中携带MoVo结合基序的基因的转录因子。
We previously described two isoforms (MOVO-A and -B) of a novel zinc finger protein MOVO, a mouse homologue of Drosophila Ovo protein. Here, we isolated cDNA encoding the third isoform MOVO-C, which had a transactivation domain and zinc finger domain, but lacked an N-terminal potential repression domain that was present in MOVO-A. Three isoform mRNAs were expressed highly in mouse testis and also in the ovary at lower levels. The structural analyses of the isolated Movo gene and mRNAs demonstrated that three different Movo transcripts were differentially processed to generate three isoforms. Major mRNA species encoded MOVO-B with a zinc finger domain alone, and minor mRNA species encoded MOVO-A (potential repressor) and MOVO-C (potential activator). To assign MOVO to a transcriptional factor, we characterized DNA-binding and transactivation properties. Random oligonucleotide selection, electrophoretic mobility shift assay and footprinting indicated that MOVO bound to the sequence, 5′-G(G/C/T)GGGGG-3′. These motifs were found in the 5′-flanking regions of Movo and other testis-specific genes. Nuclear proteins binding to this motif were detected in mouse testis, and the expression of MOVO mRNA was restricted in spermatocytes. The luciferase assay demonstrated that MOVO-C activated Movo promoter and MOVO-A repressed it, but MOVO-B had no effects. Mutated MOVO-binding motifs in the Movo promoter reduced the luciferase activity. All the isoforms had no effects on SV40 promoter without MOVO-binding motifs. MOVO-A partially rescued oogenesis of a Drosophila ovo mutant. These results suggest that MOVO isoforms are transcription factors to regulate genes carrying the MOVO-binding motifs in the testis.