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Human T-lymphotropic virus-1 visualized at the virological synapse by electron tomography.

Majorovits E, Nejmeddine M, Tanaka Y, Taylor GP, Fuller SD, Bangham CR - PLoS ONE (2008)

Bottom Line: The synaptic clefts are surrounded by the tightly apposed plasma membranes of the two cells.HTLV-1 virions can contact the recipient cell membrane before detaching from the infected cell.The results show that the HTLV-1 virological synapse that forms spontaneously between lymphocytes of HTLV-1 infected individuals allows direct cell-cell transmission of the virus by triggered, directional release of enveloped HTLV-1 particles into confined intercellular spaces.

View Article: PubMed Central - PubMed

Affiliation: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom.

ABSTRACT
Human T-lymphotropic virus 1 (HTLV-1) is transmitted directly between cells via an organized cell-cell contact called a virological synapse (VS). The VS has been studied by light microscopy, but the ultrastructure of the VS and the nature of the transmitted viral particle have remained unknown. Cell-free enveloped virions of HTLV-1 are undetectable in the serum of individuals infected with the human T-lymphotropic virus 1 (HTLV-1) and during in vitro culture of naturally infected lymphocytes. However, the viral envelope protein is required for infectivity of HTLV-1, suggesting that complete, enveloped HTLV-1 virions are transferred across the synapse. Here, we use electron tomography combined with immunostaining of viral protein to demonstrate the presence of enveloped HTLV-1 particles within the VS formed between naturally infected lymphocytes. We show in 3D that HTLV-1 particles can be detected in multiple synaptic clefts at different locations simultaneously within the same VS. The synaptic clefts are surrounded by the tightly apposed plasma membranes of the two cells. HTLV-1 virions can contact the recipient cell membrane before detaching from the infected cell. The results show that the HTLV-1 virological synapse that forms spontaneously between lymphocytes of HTLV-1 infected individuals allows direct cell-cell transmission of the virus by triggered, directional release of enveloped HTLV-1 particles into confined intercellular spaces.

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VS membrane-membrane distances and HTLV-1 virion diameter.A, B Tomogram slices showing close membrane apposition in the HTLV-1 virological synapse (VS) (black arrowheads): A, VS formed between an HTLV-1 infected MS9 cell as a donor and Jurkat cell as target. B, VS formed between a CD4+ T-cell naturally infected with HTLV-1 (PBMC) and an autologous uninfected CD4+ as a target cell. These cells were stained against HTLV-1 Gag p19 matrix protein with a specific monoclonal antibody (GIN7). The microtubule organizing centre is polarized to the synapse as revealed by one of the two centrioles (black arrow). C, The spacings between those areas of the conjugated cell membranes that constitute the VS were measured in three VS between naturally HTLV-1 infected CD4+ T-cell (PBMC) and autologous uninfected CD4+ as target cells: each distribution shows a peak around 20 nm. D, Tomogram slice showing a large number of virus particles (examples indicated by black arrows) that budded from an MS9 cell into the extra-cellular space. E, Tomogram slice showing virions (two examples indicated by black arrows) that budded from a naturally HTLV-1 infected CD4+ T-cell (PBMC) into the synaptic cleft. Most of these virions are not attached to a cell membrane. F, Distribution of diameter of HTLV-1 virions from MS9 cells (dark grey) and naturally infected CD4+ T cells (light grey): the distributions are slightly different with a median of 124 nm and 108 nm, respectively. Scale bars: A 1000 nm, B,D,E 500 nm.
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pone-0002251-g003: VS membrane-membrane distances and HTLV-1 virion diameter.A, B Tomogram slices showing close membrane apposition in the HTLV-1 virological synapse (VS) (black arrowheads): A, VS formed between an HTLV-1 infected MS9 cell as a donor and Jurkat cell as target. B, VS formed between a CD4+ T-cell naturally infected with HTLV-1 (PBMC) and an autologous uninfected CD4+ as a target cell. These cells were stained against HTLV-1 Gag p19 matrix protein with a specific monoclonal antibody (GIN7). The microtubule organizing centre is polarized to the synapse as revealed by one of the two centrioles (black arrow). C, The spacings between those areas of the conjugated cell membranes that constitute the VS were measured in three VS between naturally HTLV-1 infected CD4+ T-cell (PBMC) and autologous uninfected CD4+ as target cells: each distribution shows a peak around 20 nm. D, Tomogram slice showing a large number of virus particles (examples indicated by black arrows) that budded from an MS9 cell into the extra-cellular space. E, Tomogram slice showing virions (two examples indicated by black arrows) that budded from a naturally HTLV-1 infected CD4+ T-cell (PBMC) into the synaptic cleft. Most of these virions are not attached to a cell membrane. F, Distribution of diameter of HTLV-1 virions from MS9 cells (dark grey) and naturally infected CD4+ T cells (light grey): the distributions are slightly different with a median of 124 nm and 108 nm, respectively. Scale bars: A 1000 nm, B,D,E 500 nm.

Mentions: We used the tomographic data to measure the intermembrane spacing in the VS and the diameter of HTLV-1 particles. Areas of continuous and close apposition of the plasma membranes were present at the cell-cell contact (Figure 3A,B). The membrane spacing in the adhesion area in three synapses showed a distribution with a strong peak at ∼20 nm and an averaged median value of 25.7 nm (Figure 3C). HTLV-1 particles derived from naturally infected CD4+ T cells (N = 44) had a mean diameter of 105 nm (range 62–173 nm) and a median of 108 nm (Figure 3E,F). Particles derived from the MS9 cell line (N = 151) had a mean diameter of 126 nm (range 46–246 nm) and a median of 124 nm (Figure 3D,F).


Human T-lymphotropic virus-1 visualized at the virological synapse by electron tomography.

Majorovits E, Nejmeddine M, Tanaka Y, Taylor GP, Fuller SD, Bangham CR - PLoS ONE (2008)

VS membrane-membrane distances and HTLV-1 virion diameter.A, B Tomogram slices showing close membrane apposition in the HTLV-1 virological synapse (VS) (black arrowheads): A, VS formed between an HTLV-1 infected MS9 cell as a donor and Jurkat cell as target. B, VS formed between a CD4+ T-cell naturally infected with HTLV-1 (PBMC) and an autologous uninfected CD4+ as a target cell. These cells were stained against HTLV-1 Gag p19 matrix protein with a specific monoclonal antibody (GIN7). The microtubule organizing centre is polarized to the synapse as revealed by one of the two centrioles (black arrow). C, The spacings between those areas of the conjugated cell membranes that constitute the VS were measured in three VS between naturally HTLV-1 infected CD4+ T-cell (PBMC) and autologous uninfected CD4+ as target cells: each distribution shows a peak around 20 nm. D, Tomogram slice showing a large number of virus particles (examples indicated by black arrows) that budded from an MS9 cell into the extra-cellular space. E, Tomogram slice showing virions (two examples indicated by black arrows) that budded from a naturally HTLV-1 infected CD4+ T-cell (PBMC) into the synaptic cleft. Most of these virions are not attached to a cell membrane. F, Distribution of diameter of HTLV-1 virions from MS9 cells (dark grey) and naturally infected CD4+ T cells (light grey): the distributions are slightly different with a median of 124 nm and 108 nm, respectively. Scale bars: A 1000 nm, B,D,E 500 nm.
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pone-0002251-g003: VS membrane-membrane distances and HTLV-1 virion diameter.A, B Tomogram slices showing close membrane apposition in the HTLV-1 virological synapse (VS) (black arrowheads): A, VS formed between an HTLV-1 infected MS9 cell as a donor and Jurkat cell as target. B, VS formed between a CD4+ T-cell naturally infected with HTLV-1 (PBMC) and an autologous uninfected CD4+ as a target cell. These cells were stained against HTLV-1 Gag p19 matrix protein with a specific monoclonal antibody (GIN7). The microtubule organizing centre is polarized to the synapse as revealed by one of the two centrioles (black arrow). C, The spacings between those areas of the conjugated cell membranes that constitute the VS were measured in three VS between naturally HTLV-1 infected CD4+ T-cell (PBMC) and autologous uninfected CD4+ as target cells: each distribution shows a peak around 20 nm. D, Tomogram slice showing a large number of virus particles (examples indicated by black arrows) that budded from an MS9 cell into the extra-cellular space. E, Tomogram slice showing virions (two examples indicated by black arrows) that budded from a naturally HTLV-1 infected CD4+ T-cell (PBMC) into the synaptic cleft. Most of these virions are not attached to a cell membrane. F, Distribution of diameter of HTLV-1 virions from MS9 cells (dark grey) and naturally infected CD4+ T cells (light grey): the distributions are slightly different with a median of 124 nm and 108 nm, respectively. Scale bars: A 1000 nm, B,D,E 500 nm.
Mentions: We used the tomographic data to measure the intermembrane spacing in the VS and the diameter of HTLV-1 particles. Areas of continuous and close apposition of the plasma membranes were present at the cell-cell contact (Figure 3A,B). The membrane spacing in the adhesion area in three synapses showed a distribution with a strong peak at ∼20 nm and an averaged median value of 25.7 nm (Figure 3C). HTLV-1 particles derived from naturally infected CD4+ T cells (N = 44) had a mean diameter of 105 nm (range 62–173 nm) and a median of 108 nm (Figure 3E,F). Particles derived from the MS9 cell line (N = 151) had a mean diameter of 126 nm (range 46–246 nm) and a median of 124 nm (Figure 3D,F).

Bottom Line: The synaptic clefts are surrounded by the tightly apposed plasma membranes of the two cells.HTLV-1 virions can contact the recipient cell membrane before detaching from the infected cell.The results show that the HTLV-1 virological synapse that forms spontaneously between lymphocytes of HTLV-1 infected individuals allows direct cell-cell transmission of the virus by triggered, directional release of enveloped HTLV-1 particles into confined intercellular spaces.

View Article: PubMed Central - PubMed

Affiliation: Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom.

ABSTRACT
Human T-lymphotropic virus 1 (HTLV-1) is transmitted directly between cells via an organized cell-cell contact called a virological synapse (VS). The VS has been studied by light microscopy, but the ultrastructure of the VS and the nature of the transmitted viral particle have remained unknown. Cell-free enveloped virions of HTLV-1 are undetectable in the serum of individuals infected with the human T-lymphotropic virus 1 (HTLV-1) and during in vitro culture of naturally infected lymphocytes. However, the viral envelope protein is required for infectivity of HTLV-1, suggesting that complete, enveloped HTLV-1 virions are transferred across the synapse. Here, we use electron tomography combined with immunostaining of viral protein to demonstrate the presence of enveloped HTLV-1 particles within the VS formed between naturally infected lymphocytes. We show in 3D that HTLV-1 particles can be detected in multiple synaptic clefts at different locations simultaneously within the same VS. The synaptic clefts are surrounded by the tightly apposed plasma membranes of the two cells. HTLV-1 virions can contact the recipient cell membrane before detaching from the infected cell. The results show that the HTLV-1 virological synapse that forms spontaneously between lymphocytes of HTLV-1 infected individuals allows direct cell-cell transmission of the virus by triggered, directional release of enveloped HTLV-1 particles into confined intercellular spaces.

Show MeSH
Related in: MedlinePlus