scholarly journals Association of Potent Human Antiviral Cytidine Deaminases with 7SL RNA and Viral RNP in HIV-1 Virions

2010 ◽  
Vol 84 (24) ◽  
pp. 12903-12913 ◽  
Author(s):  
Wenyan Zhang ◽  
Juan Du ◽  
Kevin Yu ◽  
Tao Wang ◽  
Xiong Yong ◽  
...  
Keyword(s):  
Rna Binding ◽  
Signal Recognition Particle ◽  
Viral Inhibition ◽  
Cytidine Deaminases ◽  
Amino Terminal ◽  
7Sl Rna ◽  
Rnp Complex ◽  
Rna Interaction ◽  
Anti Hiv ◽  
Hiv 1

ABSTRACT 7SL RNA promotes the formation of the signal recognition particle that targets secretory and membrane proteins to the endoplasmic reticulum. 7SL RNA is also selectively packaged by many retroviruses, including HIV-1. Here, we demonstrate that 7SL RNA is an integral component of the viral ribonucleoprotein (RNP) complex containing Gag, viral genomic RNA, and \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathbf{tRNA}_{3}^{Lys}\) \end{document} . Only the potent anti-HIV-1 cytidine deaminases can bind to 7SL RNA and target to HIV-1 RNP. A conserved motif in the amino-terminal region of A3G is important for 7SL RNA interaction. The weak anti-HIV-1 A3C did not interact with 7SL RNA and failed to target to viral RNPs, despite efficient virion packaging. However, a chimeric construct of A3C plus the 7SL-binding amino terminus of A3G did target to viral RNPs and showed enhanced anti-HIV-1 activity. 7SL RNA binding is a conserved feature of human anti-HIV-1 cytidine deaminases. Thus, potent anti-HIV-1 cytidine deaminases have evolved to possess a unique RNA-binding ability for precise HIV-1 targeting and viral inhibition.

scholarly journals Human Immunodeficiency Virus Type 1 Vif Protein Is an Integral Component of an mRNP Complex of Viral RNA and Could Be Involved in the Viral RNA Folding and Packaging Process

2000 ◽  
Vol 74 (18) ◽  
pp. 8252-8261 ◽  
Author(s):  
Hui Zhang ◽  
Roger J. Pomerantz ◽  
Geethanjali Dornadula ◽  
Yong Sun
Keyword(s):  
Human Immunodeficiency Virus ◽  
Rna Binding ◽  
Viral Rna ◽  
Immunodeficiency Virus ◽  
Mrnp Complex ◽  
Rna Interaction ◽  
Vif Protein ◽  
Hiv 1

ABSTRACT Virion infectivity factor (Vif) is a protein encoded by human immunodeficiency virus types 1 and 2 (HIV-1 and -2) and simian immunodeficiency virus, plus other lentiviruses, and is essential for viral replication either in vivo or in culture for nonpermissive cells such as peripheral blood lymphoid cells, macrophages, and H9 T cells. Defects in the vif gene affect virion morphology and reverse transcription but not the expression of viral components. It has been shown that Vif colocalizes with Gag in cells and Vif binds to the NCp7 domain of Gag in vitro. However, it seems that Vif is not specifically packaged into virions. The molecular mechanism(s) for Vif remains unknown. In this report, we demonstrate that HIV-1 Vif is an RNA-binding protein and specifically binds to HIV-1 genomic RNA in vitro. Further, Vif binds to HIV-1 RNA in the cytoplasm of virus-producing cells to form a 40S mRNP complex. Coimmunoprecipitation and in vivo UV cross-linking assays indicated that Vif directly interact with HIV-1 RNA in the virus-producing cells. Vif-RNA binding could be displaced by Gag-RNA binding, suggesting that Vif protein in the mRNP complex may mediate viral RNA interaction with HIV-1 Gag precursors. Furthermore, we have demonstrated that these Vif mutants that lose the RNA binding activity in vitro do not supportvif-deficient HIV-1 replication in H9 T cells, suggesting that the RNA binding capacity of Vif is important for its function. Further studies regarding Vif-RNA interaction in virus-producing cells will be important for studying the function of Vif in the HIV-1 life cycle.

scholarly journals 7SL RNA Mediates Virion Packaging of the Antiviral Cytidine Deaminase APOBEC3G

2007 ◽  
Vol 81 (23) ◽  
pp. 13112-13124 ◽  
Author(s):  
Tao Wang ◽  
Chunjuan Tian ◽  
Wenyan Zhang ◽  
Kun Luo ◽  
Phuong Thi Nguyen Sarkis ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Rna Binding ◽  
Cytidine Deaminase ◽  
Rna Packaging ◽  
Cellular Functions ◽  
7Sl Rna ◽  
Immunodeficiency Virus ◽  
Antiviral Function ◽  
Pol Iii ◽  
Hiv 1

ABSTRACT Cytidine deaminase APOBEC3G (A3G) has broad antiviral activity against diverse retroviruses and/or retrotransposons, and its antiviral functions are believed to rely on its encapsidation into virions in an RNA-dependent fashion. However, the cofactors of A3G virion packaging have not yet been identified. We demonstrate here that A3G selectively interacts with certain polymerase III (Pol III)-derived RNAs, including Y3 and 7SL RNAs. Among A3G-binding Pol III-derived RNAs, 7SL RNA was preferentially packaged into human immunodeficiency virus type 1 (HIV-1) particles. Efficient packaging of 7SL RNA, as well as A3G, was mediated by the RNA-binding nucleocapsid domain of HIV-1 Gag. A3G mutants that had reduced 7SL RNA binding but maintained wild-type levels of mRNA and tRNA binding were packaged poorly and had impaired antiviral activity. Reducing 7SL RNA packaging by overexpression of SRP19 proteins inhibited 7SL RNA and A3G virion packaging and impaired its antiviral function. Thus, 7SL RNA that is encapsidated into diverse retroviruses is a key cofactor of the antiviral A3G. This selective interaction of A3G with certain Pol III-derived RNAs raises the question of whether A3G and its cofactors may have as-yet-unidentified cellular functions.

scholarly journals Trimer Enhancement Mutation Effects on HIV-1 Matrix Protein Binding Activities

2016 ◽  
Vol 90 (12) ◽  
pp. 5657-5664 ◽  
Author(s):  
Ayna Alfadhli ◽  
Andrew Mack ◽  
Christopher Ritchie ◽  
Isabel Cylinder ◽  
Logan Harper ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Matrix Protein ◽  
Rna Binding ◽  
Mutant Proteins ◽  
Virus Particles ◽  
Amino Terminal ◽  
Virion Incorporation ◽  
Multivalent Binding ◽  
Env Protein ◽  
Hiv 1

ABSTRACTThe HIV-1 matrix (MA) protein is the amino-terminal domain of the HIV-1 precursor Gag (Pr55Gag) protein. MA binds to membranes and RNAs, helps transport Pr55Gag proteins to virus assembly sites at the plasma membranes of infected cells, and facilitates the incorporation of HIV-1 envelope (Env) proteins into virions by virtue of an interaction with the Env protein cytoplasmic tails (CTs). MA has been shown to crystallize as a trimer and to organize on membranes in hexamer lattices. MA mutations that localize to residues near the ends of trimer spokes have been observed to impair Env protein assembly into virus particles, and several of these are suppressed by the 62QR mutation at the hubs of trimer interfaces. We have examined the binding activities of wild-type (WT) MA and 62QR MA variants and found that the 62QR mutation stabilized MA trimers but did not alter the way MA proteins organized on membranes. Relative to WT MA, the 62QR protein showed small effects on membrane and RNA binding. However, 62QR proteins bound significantly better to Env CTs than their WT counterparts, and CT binding efficiencies correlated with trimerization efficiencies. Our data suggest a model in which multivalent binding of trimeric HIV-1 Env proteins to MA trimers contributes to the process of Env virion incorporation.IMPORTANCEThe HIV-1 Env proteins assemble as trimers, and incorporation of the proteins into virus particles requires an interaction of Env CT domains with the MA domains of the viral precursor Gag proteins. Despite this knowledge, little is known about the mechanisms by which MA facilitates the virion incorporation of Env proteins. To help elucidate this process, we examined the binding activities of an MA mutant that stabilizes MA trimers. We found that the mutant proteins organized similarly to WT proteins on membranes, and that mutant and WT proteins revealed only slight differences in their binding to RNAs or lipids. However, the mutant proteins showed better binding to Env CTs than the WT proteins, and CT binding correlated with MA trimerization. Our results suggest that multivalent binding of trimeric HIV-1 Env proteins to MA trimers contributes to the process of Env virion incorporation.

scholarly journals Targeting Tat-TAR RNA Interaction for HIV-1 Inhibition

2021 ◽  
Author(s):  
Awadh Alanazi ◽  
Andrey Ivanov ◽  
Namita Kumari ◽  
Xionghao Lin ◽  
Songping Wang ◽  
...  
Keyword(s):  
Selective Inhibition ◽  
Host Factors ◽  
Tat Protein ◽  
Toxic Compounds ◽  
Cyclin T1 ◽  
Inhibitory Compounds ◽  
Tar Rna ◽  
Rna Interaction ◽  
Anti Hiv ◽  
Hiv 1

HIV-1 Tat protein interacts with TAR RNA and recruits CDK9/cyclin T1 and other host factors to induce HIV-1 transcription. Thus Tat-TAR RNA interaction, which is unique for HIV-1, represents an attractive target for anti-HIV-1 therapeutics. To target Tat-TAR RNA interaction, we used a crystal structure of TAR RNA with acetylpromazine bound to the bulge of TAR RNA, to dock compounds from Enamine database containing 1.6 million individual compounds. Docking identified 173 compounds that were analyzed for the inhibition of HIV-1 infection. Top ten inhibitory compounds with IC50 ≤ 6 µM were selected and the three least toxic compounds, T6780107 (IC50=2.97 μM), T0516-4834 (IC50=0.2 μM) and T5628834 (IC50=3.46 μM), were further tested for HIV-1 transcription inhibition. Only T0516-4834 compound showed selective inhibition of Tat-induced HIV-1 transcription, whereas T6780107 compound inhibited equally basal and Tat-induced transcription and T5628834 compound only inhibited basal HIV-1 transcription. The T0516-4834 compound also showed strongest inhibition of HIV-1 gag RNA expression and p24 production in CEM T cells infected with HIV-1 IIIB. Of the three compounds, only the T0516-4834 compound disrupted Tat-TAR RNA interaction indicating that it might target TAR RNA. Also, of the three tested compounds, T5628834 but not T6780107 or T0516-4834 disrupted Tat-CDK9/cyclin T1 interaction. Taken together, our study identified novel compound T0516-4834 that disrupted Tat-TAR RNA interaction and inhibited Tat-induced transcription and HIV-1 infection suggesting that this compound might serve as a new lead for anti-HIV-1 therapeutics.

scholarly journals A unique conformational escape reaction of HIV-1 against an allosteric integrase inhibitor

2019 ◽  
Author(s):  
Tomofumi Nakamura ◽  
Teruya Nakamura ◽  
Masayuki Amano ◽  
Toshikazu Miyakawa ◽  
Yuriko Yamagata ◽  
...  
Keyword(s):  
Rna Binding ◽  
Integrase Inhibitor ◽  
Catalytic Core ◽  
Viral Particles ◽  
Dimerization Interface ◽  
Conformational Alteration ◽  
Escape Reaction ◽  
Viral Maturation ◽  
Anti Hiv ◽  
Hiv 1

AbstractHIV-1 integrase (IN) contributes to HIV-1 RNA binding, which is required for viral maturation. Non-catalytic site integrase inhibitors (NCINIs) have been developed as allosteric IN inhibitors, which perform anti-HIV-1 activity by disrupting IN multimerization. Here, we show that IN undergoes a novel conformational alteration to escape from NCINIs. We observed that NCINI-resistant HIV-1 variants have accumulated amino acid (AA) mutations in the IN-encoding region. We employed HPLC and thermal stability assays to show that the AA mutations affect the folding and dimerization interface of the IN catalytic core domains, resulting in severely decreased multimerization of full-length IN proteins (IN under-multimerization). The under-multimerization of IN was finally restored by HIV-1 RNA in the viral particles. Our study demonstrates that HIV-1 countervails NCINIs by IN under-multimerization as a novel escape mechanism. Our findings provide information on the understanding of IN multimerization and influence the development of unique anti-HIV-1 strategies.

scholarly journals Flexibility in Nucleic Acid Binding Is Central to APOBEC3H Antiviral Activity

2019 ◽  
Vol 93 (24) ◽  
Author(s):  
Jennifer A. Bohn ◽  
Justin DaSilva ◽  
Siarhei Kharytonchyk ◽  
Maria Mercedes ◽  
Jennifer Vosters ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nucleic Acid ◽  
Antiviral Activity ◽  
Reverse Transcription ◽  
Rna Binding ◽  
Viral Dna ◽  
Dna Substrates ◽  
Apobec3 Proteins ◽  
Rna Interaction ◽  
Hiv 1

ABSTRACT APOBEC3 proteins APOBEC3F (A3F), APOBEC3G (A3G), and APOBEC3H (A3H) are host restriction factors that inhibit HIV-1 through DNA cytidine deaminase-dependent and -independent mechanisms and have either one (A3H) or two (A3F and A3G) zinc-binding domains. A3H antiviral activity encompasses multiple molecular functions, all of which depend on recognition of RNA or DNA. A3H crystal structures revealed an unusual interaction with RNA wherein an RNA duplex mediates dimerization of two A3H proteins. In this study, we sought to determine the importance of RNA-binding amino acids in the antiviral and biochemical properties of A3H. We show that the wild-type A3H-RNA interaction is essential for A3H antiviral activity and for two deaminase-independent processes: encapsidation into viral particles and inhibition of reverse transcription. Furthermore, an extensive mutagenesis campaign revealed distinct roles for two groups of amino acids at the RNA binding interface. C-terminal helix residues exclusively bind RNA, and loop 1 residues play a dual role in recognition of DNA substrates and in RNA binding. Weakening the interface between A3H and RNA allows DNA substrates to bind with greater affinity and enhances deamination rates, suggesting that RNA binding must be disrupted to accommodate DNA. Intriguingly, we demonstrate that A3H can deaminate overhanging DNA strands of RNA/DNA heteroduplexes, which are early intermediates during reverse transcription and may represent natural A3H substrates. Overall, we present a mechanistic model of A3H restriction and a step-by-step elucidation of the roles of RNA-binding residues in A3H activity, particle incorporation, inhibition of reverse transcriptase inhibition, and DNA cytidine deamination. IMPORTANCE APOBEC3 proteins are host factors that protect the integrity of the host genome by inhibiting retroelements as well as retroviruses, such as HIV-1. To do this, the APOBEC3H protein has evolved unique interactions with structured RNAs. Here, we studied the importance of these interactions in driving antiviral activity of APOBEC3H. Our results provide a clear picture of how RNA binding drives the ability of APOBEC3H to infiltrate new viruses and prevent synthesis of viral DNA. We also explore how RNA binding by APOBEC3H influences recognition and deamination of viral DNA and describe two possible routes by which APOBEC3H might hypermutate the HIV-1 genome. These results highlight how one protein can sense many nucleic acid species for a variety of antiviral activities.

scholarly journals Genetic Analysis of the Localization of APOBEC3F to Human Immunodeficiency Virus Type 1 Virion Cores

2014 ◽  
Vol 89 (4) ◽  
pp. 2415-2424 ◽  
Author(s):  
John P. Donahue ◽  
Rebecca T. Levinson ◽  
Jonathan H. Sheehan ◽  
Lorraine Sutton ◽  
Harry E. Taylor ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Cytidine Deaminase ◽  
Cytidine Deaminases ◽  
Terminal Domain ◽  
Immunodeficiency Virus ◽  
Anti Hiv ◽  
Hiv 1

ABSTRACTMembers of the APOBEC3 family of cytidine deaminases vary in their proportions of a virion-incorporated enzyme that is localized to mature retrovirus cores. We reported previously that APOBEC3F (A3F) was highly localized into mature human immunodeficiency virus type 1 (HIV-1) cores and identified that L306 in the C-terminal cytidine deaminase (CD) domain contributed to its core localization (C. Song, L. Sutton, M. Johnson, R. D'Aquila, J. Donahue, J Biol Chem287:16965–16974, 2012,http://dx.doi.org/10.1074/jbc.M111.310839). We have now determined an additional genetic determinant(s) for A3F localization to HIV-1 cores. We found that one pair of leucines in each of A3F's C-terminal and N-terminal CD domains jointly determined the degree of localization of A3F into HIV-1 virion cores. These are A3F L306/L368 (C-terminal domain) and A3F L122/L184 (N-terminal domain). Alterations to one of these specific leucine residues in either of the two A3F CD domains (A3F L368A, L122A, and L184A) decreased core localization and diminished HIV restriction without changing virion packaging. Furthermore, double mutants in these leucine residues in each of A3F's two CD domains (A3F L368A plus L184A or A3F L368A plus L122A) still were packaged into virions but completely lost core localization and anti-HIV activity. HIV virion core localization of A3F is genetically separable from its virion packaging, and anti-HIV activity requires some core localization.IMPORTANCESpecific leucine-leucine interactions are identified as necessary for A3F's core localization and anti-HIV activity but not for its packaging into virions. Understanding these signals may lead to novel strategies to enhance core localization that may augment effects of A3F against HIV and perhaps of other A3s against retroviruses, parvoviruses, and hepatitis B virus.

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