In-Vitro Subtype-Specific Modulation of HIV-1 Trans-Activator of Transcription (Tat) on RNAi Silencing Suppressor Activity and Cell Death
Abstract
:1. Introduction
2. Materials and Methods
2.1. Plasmids and Antibodies
2.2. Cell Culture and Transfection
2.3. Immuno-Blotting
2.4. Reverse Transcriptase-PCR
2.5. RNAi Silencing Suppressor Assay
2.6. Propidium Iodide (PI) Staining
2.7. Statistical Analysis
2.8. Data Availability
3. Results
3.1. Expression of Tat Subtypes and Variants at Protein and RNA Levels
3.2. RNAi Silencing Suppression (RSS) Activity of Tat Subtypes and Variants
3.3. Induction of Cell Death by Tat Subtypes and Variants
4. Discussion
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
- Ronsard, L.; Lata, S.; Singh, J.; Ramachandran, V.G.; Das, S.; Banerjea, A.C. Molecular and genetic characterization of natural HIV-1 Tat Exon-1 variants from North India and their functional implications. PLoS ONE 2014, 9, e85452. [Google Scholar] [CrossRef]
- Bbosa, N.; Kaleebu, P.; Ssemwanga, D. HIV subtype diversity worldwide. Curr. Opin. HIV AIDS 2019, 14, 153–160. [Google Scholar] [CrossRef]
- Cuevas, J.M.; Geller, R.; Garijo, R.; Lopez-Aldeguer, J.; Sanjuan, R. Extremely High Mutation Rate of HIV-1 In Vivo. PLoS Biol. 2015, 13, e1002251. [Google Scholar] [CrossRef]
- Ronsard, L.; Sood, V.; Yousif, A.S.; Ramesh, J.; Shankar, V.; Das, J.; Sumi, N.; Rai, T.; Mohankumar, K.; Sridharan, S.; et al. Genetic Polymorphisms in the Open Reading Frame of the CCR5 gene From HIV-1 Seronegative and Seropositive Individuals From National Capital Regions of India. Sci. Rep. 2019, 9, 7594. [Google Scholar] [CrossRef]
- Kaleebu, P.; Ross, A.; Morgan, D.; Yirrell, D.; Oram, J.; Rutebemberwa, A.; Lyagoba, F.; Hamilton, L.; Biryahwaho, B.; Whitworth, J. Relationship between HIV-1 Env subtypes A and D and disease progression in a rural Ugandan cohort. AIDS 2001, 15, 293–299. [Google Scholar] [CrossRef] [Green Version]
- Senkaali, D.; Muwonge, R.; Morgan, D.; Yirrell, D.; Whitworth, J.; Kaleebu, P. The relationship between HIV type 1 disease progression and V3 serotype in a rural Ugandan cohort. AIDS Res. Hum. Retrovir. 2004, 20, 932–937. [Google Scholar] [CrossRef]
- Ronsard, L.; Ganguli, N.; Singh, V.K.; Mohankumar, K.; Rai, T.; Sridharan, S.; Pajaniradje, S.; Kumar, B.; Rai, D.; Chaudhuri, S.; et al. Impact of Genetic Variations in HIV-1 Tat on LTR-Mediated Transcription via TAR RNA Interaction. Front. Microbiol. 2017, 8, 706. [Google Scholar] [CrossRef] [Green Version]
- Ronsard, L.; Rai, T.; Rai, D.; Ramachandran, V.G.; Banerjea, A.C. In silico Analyses of Subtype Specific HIV-1 Tat-TAR RNA Interaction Reveals the Structural Determinants for Viral Activity. Front. Microbiol. 2017, 8, 1467. [Google Scholar] [CrossRef]
- Raja, R.; Ronsard, L.; Lata, S.; Trivedi, S.; Banerjea, A.C. HIV-1 Tat potently stabilises Mdm2 and enhances viral replication. Biochem. J. 2017, 474, 2449–2464. [Google Scholar] [CrossRef]
- Romani, B.; Engelbrecht, S.; Glashoff, R.H. Functions of Tat: The versatile protein of human immunodeficiency virus type 1. J. Gen. Virol. 2010, 91, 1–12. [Google Scholar] [CrossRef]
- Kuppuswamy, M.; Subramanian, T.; Srinivasan, A.; Chinnadurai, G. Multiple functional domains of Tat, the trans-activator of HIV-1, defined by mutational analysis. Nucleic Acids Res. 1989, 17, 3551–3561. [Google Scholar] [CrossRef]
- Dingwall, C.; Ernberg, I.; Gait, M.J.; Green, S.M.; Heaphy, S.; Karn, J.; Lowe, A.D.; Singh, M.; Skinner, M.A. HIV-1 tat protein stimulates transcription by binding to a U-rich bulge in the stem of the TAR RNA structure. EMBO J. 1990, 9, 4145–4153. [Google Scholar] [CrossRef]
- Arya, S.K.; Guo, C.; Josephs, S.F.; Wong-Staal, F. Trans-activator gene of human T-lymphotropic virus type III (HTLV-III). Science 1985, 229, 69–73. [Google Scholar] [CrossRef]
- Cullen, B.R. Trans-activation of human immunodeficiency virus occurs via a bimodal mechanism. Cell 1986, 46, 973–982. [Google Scholar] [CrossRef]
- Rice, A.P. The HIV-1 Tat Protein: Mechanism of Action and Target for HIV-1 Cure Strategies. Curr. Pharm. Des. 2017, 23, 4098–4102. [Google Scholar] [CrossRef] [Green Version]
- Roy, S.; Delling, U.; Chen, C.H.; Rosen, C.A.; Sonenberg, N. A bulge structure in HIV-1 TAR RNA is required for Tat binding and Tat-mediated trans-activation. Genes Dev. 1990, 4, 1365–1373. [Google Scholar] [CrossRef]
- Hauber, J.; Malim, M.H.; Cullen, B.R. Mutational analysis of the conserved basic domain of human immunodeficiency virus tat protein. J. Virol. 1989, 63, 1181–1187. [Google Scholar] [Green Version]
- Frankel, A.D.; Pabo, C.O. Cellular uptake of the tat protein from human immunodeficiency virus. Cell 1988, 55, 1189–1193. [Google Scholar] [CrossRef]
- Lopez-Huertas, M.R.; Callejas, S.; Abia, D.; Mateos, E.; Dopazo, A.; Alcami, J.; Coiras, M. Modifications in host cell cytoskeleton structure and function mediated by intracellular HIV-1 Tat protein are greatly dependent on the second coding exon. Nucleic Acids Res. 2010, 38, 3287–3307. [Google Scholar] [CrossRef] [Green Version]
- Qian, S.; Zhong, X.; Yu, L.; Ding, B.; de Haan, P.; Boris-Lawrie, K. HIV-1 Tat RNA silencing suppressor activity is conserved across kingdoms and counteracts translational repression of HIV-1. Proc. Nat. Acad. Sci. USA 2009, 106, 605–610. [Google Scholar] [CrossRef] [Green Version]
- Dabrowska, A.; Kim, N.; Aldovini, A. Tat-induced FOXO3a is a key mediator of apoptosis in HIV-1-infected human CD4 + T lymphocytes. J. Immunol. 2008, 181, 8460–8477. [Google Scholar] [CrossRef]
- Sidahmed, A.; Abdalla, S.; Mahmud, S.; Wilkie, B. Antiviral innate immune response of RNA interference. J. Infect. Dev. Ctries 2014, 8, 804–810. [Google Scholar] [CrossRef]
- Bivalkar-Mehla, S.; Vakharia, J.; Mehla, R.; Abreha, M.; Kanwar, J.R.; Tikoo, A.; Chauhan, A. Viral RNA silencing suppressors (RSS): Novel strategy of viruses to ablate the host RNA interference (RNAi) defense system. Virus Res. 2011, 155, 1–9. [Google Scholar] [CrossRef] [Green Version]
- De Vries, W.; Haasnoot, J.; Fouchier, R.; de Haan, P.; Berkhout, B. Differential RNA silencing suppression activity of NS1 proteins from different influenza A virus strains. J. Gen. Virol. 2009, 90, 1916–1922. [Google Scholar] [CrossRef]
- Von Eije, K.J.; Berkhout, B. RNA-interference-based gene therapy approaches to HIV type-1 treatment: Tackling the hurdles from bench to bedside. Antivir. Chem. Chemother. 2009, 19, 221–233. [Google Scholar] [CrossRef]
- Balasubramaniam, M.; Pandhare, J.; Dash, C. Are microRNAs Important Players in HIV-1 Infection? An Update. Viruses 2018, 10, 110. [Google Scholar] [CrossRef]
- Bennasser, Y.; Le, S.Y.; Benkirane, M.; Jeang, K.T. Evidence that HIV-1 encodes an siRNA and a suppressor of RNA silencing. Immunity 2005, 22, 607–619. [Google Scholar] [CrossRef]
- Kurosu, T.; Mukai, T.; Komoto, S.; Ibrahim, M.S.; Li, Y.G.; Kobayashi, T.; Tsuji, S.; Ikuta, K. Human immunodeficiency virus type 1 subtype C exhibits higher transactivation activity of Tat than subtypes B and E. Microbiol. Immunol. 2002, 46, 787–799. [Google Scholar] [CrossRef]
- Modjarrad, K.; Vermund, S.H. Effect of treating co-infections on HIV-1 viral load: A systematic review. Lancet Infect. Dis. 2010, 10, 455–463. [Google Scholar] [CrossRef]
- Badley, A.D.; Pilon, A.A.; Landay, A.; Lynch, D.H. Mechanisms of HIV-associated lymphocyte apoptosis. Blood 2000, 96, 2951–2964. [Google Scholar] [CrossRef]
- Boasso, A.; Shearer, G.M.; Chougnet, C. Immune dysregulation in human immunodeficiency virus infection: Know it, fix it, prevent it? J. Intern. Med. 2009, 265, 78–96. [Google Scholar] [CrossRef] [PubMed]
- Rayne, F.; Debaisieux, S.; Yezid, H.; Lin, Y.L.; Mettling, C.; Konate, K.; Chazal, N.; Arold, S.T.; Pugniere, M.; Sanchez, F.; et al. Phosphatidylinositol-(4,5)-bisphosphate enables efficient secretion of HIV-1 Tat by infected T-cells. EMBO J. 2010, 29, 1348–1362. [Google Scholar] [CrossRef] [PubMed]
- Ensoli, B.; Barillari, G.; Salahuddin, S.Z.; Gallo, R.C.; Wong-Staal, F. Tat protein of HIV-1 stimulates growth of cells derived from Kaposi’s sarcoma lesions of AIDS patients. Nature 1990, 345, 84–86. [Google Scholar] [CrossRef] [PubMed]
- Sood, V.; Ranjan, R.; Banerjea, A.C. Functional analysis of HIV-1 subtypes B and C HIV-1 Tat exons and RGD/QGD motifs with respect to Tat-mediated transactivation and apoptosis. AIDS 2008, 22, 1683–1685. [Google Scholar] [CrossRef]
- Campbell, G.R.; Pasquier, E.; Watkins, J.; Bourgarel-Rey, V.; Peyrot, V.; Esquieu, D.; Barbier, P.; de Mareuil, J.; Braguer, D.; Kaleebu, P.; et al. The glutamine-rich region of the HIV-1 Tat protein is involved in T-cell apoptosis. J. Biol. Chem. 2004, 279, 48197–48204. [Google Scholar] [CrossRef]
- Jia, H.; Lohr, M.; Jezequel, S.; Davis, D.; Shaikh, S.; Selwood, D.; Zachary, I. Cysteine-rich and basic domain HIV-1 Tat peptides inhibit angiogenesis and induce endothelial cell apoptosis. Biochem. Biophys. Res. Commun. 2001, 283, 469–479. [Google Scholar] [CrossRef]
- He, M.; Zhang, L.; Wang, X.; Huo, L.; Sun, L.; Feng, C.; Jing, X.; Du, D.; Liang, H.; Liu, M.; et al. Systematic Analysis of the Functions of Lysine Acetylation in the Regulation of Tat Activity. PLoS ONE 2013, 8, e67186. [Google Scholar] [CrossRef]
- Kong, W.; Tian, C.; Liu, B.; Yu, X.F. Stable expression of primary human immunodeficiency virus type 1 structural gene products by use of a noncytopathic sindbis virus vector. J. Virol. 2002, 76, 11434–11439. [Google Scholar] [CrossRef]
- Giacca, M. HIV-1 Tat, apoptosis and the mitochondria: A tubulin link? Retrovirology 2005, 2, 7. [Google Scholar] [CrossRef]
- Kim, T.A.; Avraham, H.K.; Koh, Y.H.; Jiang, S.; Park, I.W.; Avraham, S. HIV-1 Tat-mediated apoptosis in human brain microvascular endothelial cells. J. Immunol. 2003, 170, 2629–2637. [Google Scholar] [CrossRef]
- Park, I.W.; Ullrich, C.K.; Schoenberger, E.; Ganju, R.K.; Groopman, J.E. HIV-1 Tat induces microvascular endothelial apoptosis through caspase activation. J. Immunol. 2001, 167, 2766–2771. [Google Scholar] [CrossRef] [PubMed]
- Verma, S.; Ronsard, L.; Kapoor, R.; Banerjea, A.C. Genetic characterization of natural variants of Vpu from HIV-1 infected individuals from Northern India and their impact on virus release and cell death. PLoS ONE 2013, 8, e59283. [Google Scholar] [CrossRef] [PubMed]
- Ramalingam, S.; Kannangai, R.; Abraham, O.C.; Subramanian, S.; Rupali, P.; Pulimood, S.A.; Jesudason, M.V.; Sridharan, G. Investigation of apoptotic markers among human immunodeficiency virus (HIV-1) infected individuals. Indian J. Med. Res. 2008, 128, 728–733. [Google Scholar] [PubMed]
- Romani, B.; Engelbrecht, S.; Glashoff, R.H. Antiviral roles of APOBEC proteins against HIV-1 and suppression by Vif. Arch. Virol. 2009, 154, 1579–1588. [Google Scholar] [CrossRef]
- Vashistha, H.; Husain, M.; Kumar, D.; Singhal, P.C. Tubular cell HIV-1 gp120 expression induces caspase 8 activation and apoptosis. Ren. Fail. 2009, 31, 303–312. [Google Scholar] [CrossRef]
- Chen, D.; Wang, M.; Zhou, S.; Zhou, Q. HIV-1 Tat targets microtubules to induce apoptosis, a process promoted by the pro-apoptotic Bcl-2 relative Bim. EMBO J. 2002, 21, 6801–6810. [Google Scholar] [CrossRef] [Green Version]
- Li, C.J.; Friedman, D.J.; Wang, C.; Metelev, V.; Pardee, A.B. Induction of apoptosis in uninfected lymphocytes by HIV-1 Tat protein. Science 1995, 268, 429–431. [Google Scholar] [CrossRef]
- Karjee, S.; Minhas, A.; Sood, V.; Ponia, S.S.; Banerjea, A.C.; Chow, V.T.; Mukherjee, S.K.; Lal, S.K. The 7a accessory protein of severe acute respiratory syndrome coronavirus acts as an RNA silencing suppressor. J. Virol. 2010, 84, 10395–10401. [Google Scholar] [CrossRef]
- Marno, K.M.; Ogunkolade, B.W.; Pade, C.; Oliveira, N.M.; O’Sullivan, E.; McKnight, A. Novel restriction factor RNA-associated early-stage anti-viral factor (REAF) inhibits human and simian immunodeficiency viruses. Retrovirology 2014, 11, 3. [Google Scholar] [CrossRef]
- Ronsard, L.; Raja, R.; Panwar, V.; Saini, S.; Mohankumar, K.; Sridharan, S.; Padmapriya, R.; Chaudhuri, S.; Ramachandran, V.G.; Banerjea, A.C. Genetic and functional characterization of HIV-1 Vif on APOBEC3G degradation: First report of emergence of B/C recombinants from North India. Sci. Rep. 2015, 5, 15438. [Google Scholar] [CrossRef]
- Das, A.T.; Harwig, A.; Berkhout, B. The HIV-1 Tat protein has a versatile role in activating viral transcription. J. Virol. 2011, 85, 9506–9516. [Google Scholar] [CrossRef] [PubMed]
- Yeung, M.L.; Bennasser, Y.; Le, S.Y.; Jeang, K.T. siRNA, miRNA and HIV: Promises and challenges. Cell Res. 2005, 15, 935–946. [Google Scholar] [CrossRef] [PubMed]
- Yeung, M.L.; Bennasser, Y.; Myers, T.G.; Jiang, G.; Benkirane, M.; Jeang, K.T. Changes in microRNA expression profiles in HIV-1-transfected human cells. Retrovirology 2005, 2, 81. [Google Scholar] [CrossRef]
- Contreras, X.; Bennasser, Y.; Chazal, N.; Moreau, M.; Leclerc, C.; Tkaczuk, J.; Bahraoui, E. Human immunodeficiency virus type 1 Tat protein induces an intracellular calcium increase in human monocytes that requires DHP receptors: Involvement in TNF-alpha production. Virology 2005, 332, 316–328. [Google Scholar] [CrossRef] [PubMed]
- Bartz, S.R.; Emerman, M. Human immunodeficiency virus type 1 Tat induces apoptosis and increases sensitivity to apoptotic signals by up-regulating FLICE/caspase-8. J. Virol. 1999, 73, 1956–1963. [Google Scholar]
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Ronsard, L.; S. Yousif, A.; Ramesh, J.; Sumi, N.; Gorman, M.; G. Ramachandran, V.; C. Banerjea, A. In-Vitro Subtype-Specific Modulation of HIV-1 Trans-Activator of Transcription (Tat) on RNAi Silencing Suppressor Activity and Cell Death. Viruses 2019, 11, 976. https://doi.org/10.3390/v11110976
Ronsard L, S. Yousif A, Ramesh J, Sumi N, Gorman M, G. Ramachandran V, C. Banerjea A. In-Vitro Subtype-Specific Modulation of HIV-1 Trans-Activator of Transcription (Tat) on RNAi Silencing Suppressor Activity and Cell Death. Viruses. 2019; 11(11):976. https://doi.org/10.3390/v11110976
Chicago/Turabian StyleRonsard, Larance, Ashraf S. Yousif, Janani Ramesh, N. Sumi, Matthew Gorman, Vishnampettai G. Ramachandran, and Akhil C. Banerjea. 2019. "In-Vitro Subtype-Specific Modulation of HIV-1 Trans-Activator of Transcription (Tat) on RNAi Silencing Suppressor Activity and Cell Death" Viruses 11, no. 11: 976. https://doi.org/10.3390/v11110976