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Ultrastructural localization of HIV-1 RNA and core proteins. Simultaneous visualization using double immunogold
F Escaig-Haye1, V Grigoriev, I Sharova
1U 153 INSERM, St-Vincent-de-Paul Hospital, Paris, France.
Insights
This study reveals human immunodeficiency virus type 1 (HIV-1) RNA localization within infected lymphoid cells. HIV-1 RNA is found in the cytoplasm and near the cell membrane during active virus production and release.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Human immunodeficiency virus type 1 (HIV-1) infection impacts lymphoid cells.
- Understanding viral RNA distribution is crucial for comprehending HIV-1 replication and pathogenesis.
Purpose of the Study:
- To investigate the subcellular localization of HIV-1 RNA within infected lymphoid cells.
- To correlate HIV-1 RNA distribution with viral particle assembly and release.
Main Methods:
- Subcellular analysis of HIV-1 RNA using immunogold cytochemistry on Lowicryl thin sections.
- Detection of HIV-1 RNA with a biotinylated DNA probe.
- Double immunogold labeling to identify HIV-1 RNA alongside core proteins (p24, p17).
Main Results:
- HIV-1 RNA was detected in the cytoplasm, near the plasma membrane, and within extracellular virions.
- Immature and nascent virus forms showed RNA at the periphery; mature virions had RNA concentrated in the nucleoid.
- HIV-1 RNA localized to the cytoplasm, while structural proteins were found on viral formations.
Conclusions:
- Cytoplasmic HIV-1 RNA suggests genomic origin, destined for packaging into new virions.
- RNA localization near the plasma membrane coincides with viral structural protein assembly and budding.
- This subcellular distribution provides insight into HIV-1 RNA's role in the late stages of viral replication.
Abstract:
The analysis of human immunodeficiency virus type 1 (HIV-1) RNA sequences in CEM and Jurkat lymphoid cells infected with the virus has been performed at the subcellular level. Using a biotinylated DNA probe specific for HIV-1, virus RNA sequences were detected on Lowicryl thin sections after immunogold cytochemistry. The labelling observed on the cytoplasm was localized near the plasma membrane connected with extracellular cluster of virions. On free immature and nascent form of the virus the detection of HIV-1 RNA was associated with the peripheral electron-dense structure, whereas on mature form the labelling was concentrated on the central nucleoid known to be the site of the HIV-1 genomic RNA. The identification of virus RNA was also performed simultaneously with the detection of HIV-1 core protein p24 or p17 using a double immunogold labelling. Whereas the HIV-1 RNA showed again a cytoplasmic and virions localization, the structural protein was only observed on viral formations. The cytoplasmic localization of virus RNA, at the time of virus production, suggests that they are of genomic origin destined to be packaged in virions once the assembly of virus structural proteins has taken place in the plasma membrane at the viral budding site. The present molecular investigation conducted at the subcellular level provides insight into the cell periphery distribution of HIV-1 RNA observed at the light microscope as corresponding to the detection of HIV-1 infected lymphoid cells actually releasing virions.