Novel RNase H Inhibitors Blocking RNA-directed Strand Displacement DNA Synthesis by HIV-1 Reverse Transcriptase. Issue 7 (15th April 2022)

• Record Type: Journal Article
• Authors: Martín-Alonso, Samara; Kang, Dongwei; Martínez del Río, Javier; Luczkowiak, Joanna; Frutos-Beltrán, Estrella; Zhang, Lina; Cheng, Xiqiang; Liu, Xinyong; Zhan, Peng; Menéndez-Arias, Luis
• Is Part Of: Journal of molecular biology. Volume 434:Issue 7(2022)
• Publication Date: 2022-04-15
• Subjects: reverse transcriptase -- HIV -- RNase H -- RNase H inhibitors -- strand displacement; DTT dithiothreitol -- HID hydroxyisoquinoline-1, 3-dione -- HBV hepatitis B virus -- HIV human immunodeficiency virus -- HSV herpes simplex virus -- IN integrase -- LTR long terminal repeat -- MLV murine leukemia virus -- NNRTIs nonnucleoside reverse transcriptase inhibitors -- NRTIs nucleoside reverse transcriptase inhibitors -- RDDP RNA-dependent DNA polymerase -- RNase H ribonuclease H -- RT reverse transcriptase; Molecular biology -- Periodicals; Biology -- Periodicals; Biochemistry -- Periodicals; Bacteriology -- Periodicals; Molecular Biology -- Periodicals; Biochemistry -- Periodicals; Biologie moléculaire -- Périodiques; Biologie -- Périodiques; Biochimie -- Périodiques; Moleculaire biologie; Biochemistry; Biology; Molecular biology; Periodicals; 572.805
• DOI: 10.1016/j.jmb.2022.167507
• Languages: English
• ISSNs: 0022-2836

Abstract

Graphical abstract: Highlights: Retroviral RTs have a strong strand displacement activity with RNA and DNA templates. In HIV-1 RT, RNase H inactivating mutations reduce its RNA-dependent strand displacement activity. RNase H active site inhibitors produce similar effects in WT RT-catalyzed DNA synthesis reactions. Coumarin-based DNA pol/RNase H dual agents were the most effective strand displacement inhibitors. Reported findings are relevant for antiretroviral drug design and transcriptomics technologies. Abstract: In retroviruses, strand displacement DNA-dependent DNA polymerization catalyzed by the viral reverse transcriptase (RT) is required to synthesize double-stranded proviral DNA. In addition, strand displacement during RNA-dependent DNA synthesis is critical to generate high-quality cDNA for use in molecular biology and biotechnology. In this work, we show that the loss of RNase H activity due to inactivating mutations in HIV-1 RT (e.g. D443N or E478Q) has no significant effect on strand displacement while copying DNA templates, but has a large impact on DNA polymerization in reactions carried out with RNA templates. Similar effects were observed with β-thujaplicinol and other RNase H active site inhibitors, including compounds with dual activity (i.e., characterized also as inhibitors of HIV-1 integrase and/or the RT DNA polymerase). Among them, dual inhibitors of HIV-1 RT DNA polymerase/RNase H activities, containing a 7-hydroxy-6-nitro-2 H -chromen-2-one pharmacophore were found to be very potent and effective strand displacement inhibitors in RNA-dependent DNA polymerization reactions. These findings might be helpful in the development of transcriptomics technologies to obtain more uniform read coverages when copying long RNAs and for the construction of more representative libraries avoiding biases towards 5′ and 3′ ends, while providing valuable information for the development of novel antiretroviral agents.