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RNase H sequence preferences influence antisense oligonucleotide efficiency.

ABSTRACT: 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.

SUBMITTER: Kielpinski LJ

PROVIDER: S-EPMC5728404 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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RNase H sequence preferences influence antisense oligonucleotide efficiency.

Kielpinski Lukasz J LJ Hagedorn Peter H PH Lindow Morten M Vinther Jeppe J

Nucleic acids research 20171201 22

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 h ...[more]

PMID: 29126318

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Project description:BackgroundThe rapid emergence of antibiotic-resistant bacteria directly contributes to a wave of untreatable infections. The lack of new drug development is an important driver of this crisis. Most antibiotics today are small molecules that block vital processes in bacteria. To optimize such effects, the three-dimensional structure of targeted bacterial proteins is imperative, although such a task is time-consuming and tedious, impeding the development of antibiotics. The development of RNA-based therapeutics has catalyzed a new platform of antibiotics-antisense oligonucleotides (ASOs). These molecules hybridize with their target mRNAs with high specificity, knocking down or interfering with protein translation. This study aims to develop a bioinformatics pipeline to identify potent ASO targets in essential bacterial genes.MethodsThree bacterial species (P. gingivalis, H. influenzae, and S. aureus) were used to demonstrate the utility of the pipeline. Open reading frames of bacterial essential genes were downloaded from the Database of Essential Genes (DEG). After filtering for specificity and accessibility, ASO candidates were ranked based on their self-hybridization score, predicted melting temperature, and the position on the gene in an operon. Enrichment analysis was conducted on genes associated with putative potent ASOs.ResultsA total of 45,628 ASOs were generated from 348 unique essential genes in P. gingivalis. A total of 1,117 of them were considered putative. A total of 27,273 ASOs were generated from 191 unique essential genes in H. influenzae. A total of 847 of them were considered putative. A total of 175,606 ASOs were generated from 346 essential genes in S. aureus. A total of 7,061 of them were considered putative. Critical biological processes associated with these genes include translation, regulation of cell shape, cell division, and peptidoglycan biosynthetic process. Putative ASO targets generated for each bacterial species are publicly available here: https://github.com/EricSHo/AOA. The results demonstrate that our bioinformatics pipeline is useful in identifying unique and accessible ASO targets in bacterial species that post major public health issues.

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RNase H sequence preferences influence antisense oligonucleotide efficiency.

Publications

RNase H sequence preferences influence antisense oligonucleotide efficiency.

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