Ultrastructural localization of HIV-1 RNA and core proteins. Simultaneous visualization using double immunogold labelling after in situ hybridization and immunocytochemistry.
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.
Escaig-Haye
F
F
U 153 INSERM, St-Vincent-de-Paul Hospital, Paris, France.
Grigoriev
V
V
Sharova
I
I
Rudneva
V
V
Buckrinskaya
A
A
Fournier
J G
JG
D001710:Biotin; D002460:Cell Line / Q000382:microbiology; D002462:Cell Membrane / Q000382:microbiology; D015342:DNA Probes; D015497:HIV-1 / Q000737:chemistry* / Q000648:ultrastructure; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D012367:RNA, Viral / Q000032:analysis*; D014758:Viral Core Proteins / Q000032:analysis*
D015342:DNA Probes; D012367:RNA, Viral; D014758:Viral Core Proteins; D001710:Biotin