Specific Guanosines in the HIV-2 Leader RNA are Essential for Efficient Viral Genome Packaging |
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| Authors: | Chijioke N. Umunnakwe Alice Duchon Olga A. Nikolaitchik Sheikh Abdul Rahman Yang Liu Jianbo Chen Sheldon Tai Vinay K. Pathak Wei-Shau Hu |
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| Affiliation: | 1. Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute, Frederick, MD 21702, USA;2. Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute, Frederick, MD 21702, USA;1. Department of Molecular Genetics and Infection Biology, Interfaculty Institute for Genetics and Functional Genomics, Center for Functional Genomics of Microbes, University of Greifswald, D-17487 Greifswald, Germany;2. Department of Crystallography and Structural Biology, Institute of Physical Chemistry “Rocasolano”, CSIC, 28006 Madrid, Spain;3. Cellular Metabolism/Metabolomics, Institute of Biochemistry, University of Greifswald, D-17487 Greifswald, Germany;4. Present Address: Institut für Klinische Chemie und Laboratoriumsmedizin, Universitätsmedizin Greifswald, Germany;5. Present Address: Institute for Microbiology, University of Veterinary Medicine Hannover, Hannover, Germany;6. Present Address: Biochemie der Pflanzen, Heinrich-Heine-Universität Düsseldorf, Germany;7. Present Address: Structural Molecular Biology Group, Novo Nordisk Foundation Centre for Protein Research, Faculty of Health and Medical Sciences University of Copenhagen, Blegdamsvej 3-B, Copenhagen, 2200, Denmark;1. Section for Structural Biology, Department of Infectious Disease, Imperial College London, Exhibition Road, London SW7 2BB, United Kingdom;2. State Key Laboratory of Microbial Metabolism, School of Life Sciences & Biotechnology, The Joint International Research Laboratory of Metabolic & Developmental Sciences MOE, Shanghai Jiao Tong University, Shanghai, China;3. Instrumental Analysis Center, Shanghai Jiao Tong University, Shanghai, China;4. Institute of Reproductive and Developmental Biology, Department of Metabolism, Digestion and Reproduction, Imperial College London, Du Cane Road, London W12 0NN, United Kingdom;1. SCITEC-CNR, via Mario Bianco 9, 20131 Milan, Italy;2. Department of Chemistry, University of Pavia, via Taramelli 12, 27100 Pavia, Italy;1. Interdisciplinary Biological Sciences Graduate Program, Northwestern University, Evanston, IL, United States;2. Department of Chemical and Biological Engineering, Northwestern University, Evanston, IL, United States;3. US Army Combat Capabilities Development Command Chemical Biological Center, Edgewood, MD, United States;4. Molecular Biosciences Program, Weinberg College of Arts and Sciences, Northwestern University, Evanston, IL, United States;5. Center for Synthetic Biology, Northwestern University, Evanston, IL, United States |
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| Abstract: | HIV-2, a human pathogen that causes acquired immunodeficiency syndrome, is distinct from the more prevalent HIV-1 in several features including its evolutionary history and certain aspects of viral replication. Like other retroviruses, HIV-2 packages two copies of full-length viral RNA during virus assembly and efficient genome encapsidation is mediated by the viral protein Gag. We sought to define cis-acting elements in the HIV-2 genome that are important for the encapsidation of full-length RNA into viral particles. Based on previous studies of murine leukemia virus and HIV-1, we hypothesized that unpaired guanosines in the 5′ untranslated region (UTR) play an important role in Gag:RNA interactions leading to genome packaging. To test our hypothesis, we targeted 18 guanosines located in 9 sites within the HIV-2 5′ UTR and performed substitution analyses. We found that mutating as few as three guanosines significantly reduce RNA packaging efficiency. However, not all guanosines examined have the same effect; instead, a hierarchical order exists wherein a primary site, a secondary site, and three tertiary sites are identified. Additionally, there are functional overlaps in these sites and mutations of more than one site can act synergistically to cause genome packaging defects. These studies demonstrate the importance of specific guanosines in HIV-2 5′UTR in mediating genome packaging. Our results also demonstrate an interchangeable and hierarchical nature of guanosine-containing sites, which was not previously established, thereby revealing key insights into the replication mechanisms of HIV-2. |
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| Keywords: | retroviruses encapsidation RNA guanosine untranslated region |
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