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High efficiency of HIV-1 genomic RNA packaging and heterozygote formation revealed by single virion analysis

  1. Author:
    Chen, J. B.
    Nikolaitchik, O.
    Singh, J.
    Wright, A.
    Bencsics, C. E.
    Coffin, J. M.
    Ni, N.
    Lockett, S.
    Pathak, V. K.
    Hu, W. S.
  2. Author Address

    Singh, Jatinder, Wright, Andrew, Bencsics, Craig E.; Coffin, John M.] Tufts Univ, Dept Microbiol, Boston, MA 02111 USA. [Chen, Jianbo, Nikolaitchik, Olga, Ni, Na, Pathak, Vinay K.; Hu, Wei-Shau] NCI, HIV Drug Resistance Program, Frederick, MD 21702 USA. [Lockett, Stephen] Sci Applicat Int Corp, Opt Microscopy & Anal Lab, Frederick, MD 21702 USA.
    1. Year: 2009
  1. Journal: Proceedings of the National Academy of Sciences of the United States of America
    1. 106
    2. 32
    3. Pages: 13535-13540
  2. Type of Article: Article
  1. Abstract:

    A long-standing question in retrovirus biology is how RNA genomes are distributed among virions. In the studies presented in this report, we addressed this issue by directly examining HIV-1 RNAs in virions using a modified HIV-1 genome that contained recognition sites for BglG, an antitermination protein in the Escherichia coli bgl operon, which was coexpressed with a fragment of BglG RNA binding protein fused to a fluorescent protein. Our results demonstrate that the majority of virions (>90%) contain viral RNAs. We also coexpressed HIV-1 genomes containing binding sites for BglG or the bacteriophage MS2 coat protein along with 2 fluorescent protein-tagged RNA binding proteins. This method allows simultaneously labeling and discrimination of 2 different RNAs at single-RNA-detection sensitivity. Using this strategy, we obtained physical evidence that virions contain RNAs derived from different parental viruses (heterozygous virion) at ratios expected from a random distribution, and we found that this ratio can be altered by changing the dimerization sequences. Our studies of heterozygous virions also support a generally accepted but unproven assumption that most particles contain 1 dimer. This study provides answers to long-standing questions in HIV-1 biology and illustrates the power and sensitivity of the 2-RNA labeling method, which can also be adapted to analyze various issues of RNA biogenesis including the detection of different RNAs in live cell imaging.

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External Sources

  1. DOI: 10.1073/pnas.0906822106
  2. PMID: 19628694

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