Koval Anastasia P, Kramerov Dmitri A
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia.
Gene. 2009 Oct 15;446(2):75-80. doi: 10.1016/j.gene.2009.07.005. Epub 2009 Jul 17.
4.5SH RNA is a 94 nt small nuclear RNA with an unknown function. Hundreds of its genes are present in the genomes of rodents of six families including Muridae. 4.5SH RNA genes contain an internal RNA-polymerase III promoter consisting of A and B boxes. Here we studied the influence of 5'-flanking sequences on the transcription of a mouse 4.5SH RNA gene. We found that replacement of the upstream sequence can dramatically change the 4.5SH RNA gene transcription efficiency. Various DNA fragments inserted immediately upstream from 4.5SH RNA gene completely inhibited its in vitro transcription, whereas others promoted it. The shortening of the native mouse 5'-flanking sequence of 4.5SH RNA gene to 42 bp resulted in the activation of an additional illegal transcription start site in upstream region. Transcription of the 4.5SH RNA gene with various upstream sequences in transfected HeLa cells revealed the differences between the tests performed in vivo and in vitro: in whole cells, only the construct with 5'-flanking native sequence could be transcribed. Apparently, at least some regions of the native 5'-flanking sequence of 4.5SH RNA genes have been selected during evolution for high transcription activity.
4.5SH RNA是一种功能未知的94个核苷酸的小核RNA。在包括鼠科在内的六个科的啮齿动物基因组中存在数百个其基因。4.5SH RNA基因包含一个由A盒和B盒组成的内部RNA聚合酶III启动子。在此，我们研究了5'侧翼序列对小鼠4.5SH RNA基因转录的影响。我们发现上游序列的替换可显著改变4.5SH RNA基因的转录效率。紧邻4.5SH RNA基因上游插入的各种DNA片段完全抑制其体外转录，而其他片段则促进转录。将4.5SH RNA基因天然的小鼠5'侧翼序列缩短至42 bp导致上游区域额外出现一个异常转录起始位点的激活。在转染的HeLa细胞中用各种上游序列对4.5SH RNA基因进行转录，揭示了体内和体外实验之间的差异：在完整细胞中，只有带有5'侧翼天然序列的构建体能够被转录。显然，4.5SH RNA基因天然5'侧翼序列的至少一些区域在进化过程中被选择用于高效转录活性。
