Dominski Zbigniew, Yang Xiao-Cui, Purdy Matthew, Marzluff William F
Department of Biochemistry and Biophysics and Program in Molecular Biology and Biotechnology, University of North Carolina, Chapel Hill, NC 27599, USA.
Proc Natl Acad Sci U S A. 2003 Aug 5;100(16):9422-7. doi: 10.1073/pnas.1533509100. Epub 2003 Jul 18.
Base pairing between the 5' end of U7 small nuclear RNA (snRNA) and the histone downstream element (HDE) in replication-dependent histone pre-mRNAs is the key event in 3'-end processing that leads to generation of mature histone mRNAs. We have cloned the Drosophila U7 snRNA and demonstrated that it is required for histone pre-mRNA 3'-end processing in a Drosophila nuclear extract. The 71-nt Drosophila U7 snRNA is encoded by a single gene that is embedded in the direct orientation in an intron of the Eip63E gene. The U7 snRNA gene contains conserved promoter elements typical of other Drosophila snRNA genes, and the coding sequence is followed by a 3' box indicating that the Drosophila U7 snRNA gene is an independent transcription unit. Drosophila U7 snRNA contains a trimethyl-guanosine cap at the 5' end and a putative Sm-binding site similar to the unique Sm-binding site found in other U7 snRNAs. Drosophila U7 snRNA is approximately 10 nt longer than mammalian U7 snRNAs because of an extended 5' sequence and has only a limited potential to form a stem-loop structure near the 3' end. The extended 5' end of Drosophila U7 snRNA can base pair with the HDE in all five Drosophila histone pre-mRNAs. Blocking the 5' end of the U7 snRNA with a complementary oligonucleotide specifically blocks processing of a Drosophila histone pre-mRNA. Changes in the HDE that abolish or decrease processing efficiency result in a reduced ability to recruit U7 snRNA to the pre-mRNA.
U7小核RNA(snRNA)5'端与复制依赖型组蛋白前体mRNA中的组蛋白下游元件(HDE)之间的碱基配对是3'端加工过程中的关键事件,该过程导致成熟组蛋白mRNA的产生。我们克隆了果蝇U7 snRNA,并证明它是果蝇核提取物中组蛋白前体mRNA 3'端加工所必需的。71个核苷酸的果蝇U7 snRNA由单个基因编码,该基因以正向方式嵌入Eip63E基因的一个内含子中。U7 snRNA基因包含其他果蝇snRNA基因典型的保守启动子元件,编码序列后面跟着一个3'框,表明果蝇U7 snRNA基因是一个独立的转录单元。果蝇U7 snRNA在5'端含有一个三甲基鸟苷帽,以及一个类似于其他U7 snRNAs中发现的独特Sm结合位点的推定Sm结合位点。由于5'序列延长,果蝇U7 snRNA比哺乳动物U7 snRNAs大约长10个核苷酸,并且在3'端附近形成茎环结构的潜力有限。果蝇U7 snRNA延长的5'端可以与所有五种果蝇组蛋白前体mRNA中的HDE进行碱基配对。用互补寡核苷酸封闭U7 snRNA的5'端可特异性阻断果蝇组蛋白前体mRNA的加工。HDE的变化若消除或降低加工效率,会导致将U7 snRNA募集到前体mRNA上的能力降低。