Displacement of YY1 by differentiation-specific transcription factor hSkn-1a activates the P670 promoter of human papillomavirus type 16
Displacement of YY1 by differentiation-specific transcription factor hSkn-1a activates the P670 promoter of human papillomavirus type 16
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DOI：
10.1128/jvi.75.19.9302-9311.2001
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发表时间：
2001-10-01
影响因子：
5.4
通讯作者：
Kanda, T
中科院分区：
文献类型：
--
作者：
Kukimoto, I;Kanda, T
人乳头瘤病毒16型（HPV 16）P-670（E7开放阅读框中的启动子）的转录在未分化的角质形成细胞中受到抑制，但在分化时被激活。我们发现，由P-670驱动的瞬时荧光素酶表达显着增强HeLa细胞共转染表达质粒的人Skn-1a（hSkn-1a），特异性分化角质形成细胞的转录因子。单独的hSkn-1a POU结构域，其介导序列特异性DNA结合，足以激活荧光素酶的表达。电泳迁移率变动分析显示，存在两个结合位点，位点1和2，P-670的上游，这是由hSkn-1a和YY 1共享。位点1与hSkn-1a的结合强度高于位点2。与含有位点1的短DNA片段复合的YY 1被hSkn-1a取代，表明hSkn-1a与位点1的亲和力比YY 1强。通过核苷酸取代破坏结合位点，提高了荧光素酶的基础表达水平，降低了hSkn-1a的增强作用。在用环状HPV 16 DNA沿着hSkn-1a表达质粒转染的HeLa细胞中，可检测到来自P-670的转录物，这表明用报告质粒获得的结果可能模拟了真实HPV 16 DNA中P-670的调节。数据强烈表明，P-670的转录主要受到YY 1与这两个位点结合的抑制，并且hSkn-1a对YY 1的置换使P-670从抑制中释放出来。
Transcription from human papillomavirus type 16 (HPV16) P-670, a promoter in the E7 open reading frame, is repressed in undifferentiated keratinocytes but becomes activated upon differentiation. We showed that the transient luciferase expression driven by P-670 was markedly enhanced in HeLa cells cotransfected with an expression plasmid for human Skn-1a (hSkn-1a), a transcription factor specific to differentiating keratinocytes. The hSkn-1a POU domain alone, which mediates sequence-specific DNA binding, was sufficient to activate the expression of luciferase. Electrophoretic mobility shift assay revealed the presence of two binding sites, sites 1 and 2, upstream of P-670, which were shared by hSkn-1a and YY1. Site 1 bound more strongly to hSkn-1a than site 2 did. YY1 complexing with a short DNA fragment having site 1 was displaced by hSkn-1a, indicating that hSkn-1a's affinity with site 1 was stronger than YY1's. Disrupting the binding sites by nucleotide substitutions raised the basal expression level of luciferase and decreased the enhancing effect of hSkn-1a. In HeLa cells transfected with circular HPV16 DNA along with the expression plasmid for hSkn-1a, the transcript from P-670 was detectable, which indicates that the results obtained with the reporter plasmids are likely to have mimicked the regulation of P-670 in authentic HPV16 DNA. The data strongly suggest that the transcription from P-670 is repressed primarily by YY1 binding to the two sites, and the displacement of YY1 by hSkn-1a releases P-670 from the repression.