Kielpinski Lukasz J, Hagedorn Peter H, Lindow Morten, Vinther Jeppe
Roche Pharmaceutical Discovery and Early Development, Therapeutic Modalities, Roche Innovation Center Copenhagen, Fremtidsvej 3, DK-2970 Hørsholm, Denmark.
Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200 Copenhagen N, Denmark.
Nucleic Acids Res. 2017 Dec 15;45(22):12932-12944. doi: 10.1093/nar/gkx1073.
RNase H cleaves RNA in RNA-DNA duplexes. It is present in all domains of life as well as in multiple viruses and is essential for mammalian development and for human immunodeficiency virus replication. Here, we developed a sequencing-based method to measure the cleavage of thousands of different RNA-DNA duplexes and thereby comprehensively characterized the sequence preferences of HIV-1, human and Escherichia coli RNase H enzymes. We find that the catalytic domains of E. coli and human RNase H have nearly identical sequence preferences, which correlate with the efficiency of RNase H-recruiting antisense oligonucleotides. The sequences preferred by HIV-1 RNase H are distributed in the HIV genome in a way suggesting selection for efficient RNA cleavage during replication. Our findings can be used to improve the design of RNase H-recruiting antisense oligonucleotides and show that sequence preferences of HIV-1 RNase H may have shaped evolution of the viral genome and contributed to the use of tRNA-Lys3 as primer during viral replication.
核糖核酸酶H（RNase H）可切割RNA-DNA双链体中的RNA。它存在于所有生命域以及多种病毒中，对哺乳动物发育和人类免疫缺陷病毒复制至关重要。在此，我们开发了一种基于测序的方法来测量数千种不同RNA-DNA双链体的切割情况，从而全面表征了HIV-1、人类和大肠杆菌RNase H酶的序列偏好。我们发现大肠杆菌和人类RNase H的催化结构域具有几乎相同的序列偏好，这与招募RNase H的反义寡核苷酸的效率相关。HIV-1 RNase H偏好的序列在HIV基因组中的分布方式表明，在复制过程中选择了高效的RNA切割。我们的研究结果可用于改进招募RNase H的反义寡核苷酸的设计，并表明HIV-1 RNase H的序列偏好可能塑造了病毒基因组的进化，并有助于在病毒复制过程中使用tRNA-Lys3作为引物。
