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Role of the C-terminal domain of the HIV-1 glycoprotein in cell-to-cell viral transmission between T lymphocytes.

Emerson V, Haller C, Pfeiffer T, Fackler OT, Bosch V - Retrovirology (2010)

Bottom Line: We further observed that in permissive cells, which supported both routes of mutant virus transmission, viral gene expression levels, Gag processing and particle release were inherently higher than in non-permissive cells, a factor which may be significantly contributing to their permissivity phenotype.Additionally, and correlating with viral transfer efficiencies in these cell types, HIV-Gag accumulation at the virological synapse (VS) was reduced to background levels in the absence of the Env-CT in conjugates of non-permissive cells but not in permissive cells.The requirement of the Env-CT for these related processes is abrogated in permissive cells, which exhibit higher HIV gene expression levels.

View Article: PubMed Central - HTML - PubMed

Affiliation: Forschungsschwerpunkt Infektion und Krebs, F020, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.

ABSTRACT

Background: Mutant HIV (HIV-Env-Tr712) lacking the cytoplasmic tail of the viral glycoprotein (Env-CT) exhibits a cell-type specific replication phenotype such that replicative spread occurs in some T-cell lines (referred to as permissive cells) but fails to do so in most T-cell lines or in PBMCs (referred to as non-permissive cells). We aim to gain insight on the underlying requirement for the Env-CT for viral spread in non-permissive cells.

Results: We established that in comparison to HIV-Wt, both cell-free and cell-to-cell transmission of mutant HIV-Env-Tr712 from non-permissive cells were severely impaired under naturally low infection conditions. This requirement for Env-CT could be largely overcome by using saturating amounts of virus for infection. We further observed that in permissive cells, which supported both routes of mutant virus transmission, viral gene expression levels, Gag processing and particle release were inherently higher than in non-permissive cells, a factor which may be significantly contributing to their permissivity phenotype. Additionally, and correlating with viral transfer efficiencies in these cell types, HIV-Gag accumulation at the virological synapse (VS) was reduced to background levels in the absence of the Env-CT in conjugates of non-permissive cells but not in permissive cells.

Conclusions: During natural infection conditions, the HIV-Env-CT is critically required for viral transmission in cultures of non-permissive cells by both cell-free and cell-to-cell routes and is instrumental for Gag accumulation to the VS. The requirement of the Env-CT for these related processes is abrogated in permissive cells, which exhibit higher HIV gene expression levels.

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Env incorporation into Wt-HIV and HIV-Env-Tr712 virion. Producer H9 (np) cells had been weakly infected (less than 20% infection level). A. Western blot: the top part of the filter has been probed with gp120 antibodies, the middle part with gp41 mAb, Chessie 8 against the Env C-terminal tail (truncated in HIV-Env-Tr712) and the bottom part with p24 mAb. B. Average gp120 incorporation into HIV-Env-Tr712 virions in comparison to HIV-Wt (from 3 independent experiments: individual values 37%, 118%, 83%).
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Figure 3: Env incorporation into Wt-HIV and HIV-Env-Tr712 virion. Producer H9 (np) cells had been weakly infected (less than 20% infection level). A. Western blot: the top part of the filter has been probed with gp120 antibodies, the middle part with gp41 mAb, Chessie 8 against the Env C-terminal tail (truncated in HIV-Env-Tr712) and the bottom part with p24 mAb. B. Average gp120 incorporation into HIV-Env-Tr712 virions in comparison to HIV-Wt (from 3 independent experiments: individual values 37%, 118%, 83%).

Mentions: Additionally, it is likely that differences in infection levels of producer cells also account for the fact that, in contrast to our earlier report [33], others had previously reported reduction in the cell-free infectivity of HIV-Env-Tr712 virions produced in non-permissive cells [19,21]. In these latter studies, reduced infectivity had been reported to correlate with a defect in Env-Tr712 incorporation [19,21]. Thus, in order to study this here, HIV-Wt and HIV-Env-Tr712 virions were produced in weakly infected H9 (np) cells (less than 20% infected) and their protein content analysed by Western blot. Three independent experiments were evaluated by quantifying the amount of incorporated gp120 relative to viral p24 content for each virus preparation. As seen in Fig. 3A, gp120 incorporation into HIV-Env-Tr712 virions can clearly be seen and, as shown in Fig. 3B, the amount is, on average 79% of that in HIV-Wt. The reason for this discrepancy between these Env incorporation results and those previously published is presently not known. At any rate, this modest reduction in Env incorporation appears unlikely to account for the strongly reduced infectivity of cell-free HIV-Env-Tr712 virions.


Role of the C-terminal domain of the HIV-1 glycoprotein in cell-to-cell viral transmission between T lymphocytes.

Emerson V, Haller C, Pfeiffer T, Fackler OT, Bosch V - Retrovirology (2010)

Env incorporation into Wt-HIV and HIV-Env-Tr712 virion. Producer H9 (np) cells had been weakly infected (less than 20% infection level). A. Western blot: the top part of the filter has been probed with gp120 antibodies, the middle part with gp41 mAb, Chessie 8 against the Env C-terminal tail (truncated in HIV-Env-Tr712) and the bottom part with p24 mAb. B. Average gp120 incorporation into HIV-Env-Tr712 virions in comparison to HIV-Wt (from 3 independent experiments: individual values 37%, 118%, 83%).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2875203&req=5

Figure 3: Env incorporation into Wt-HIV and HIV-Env-Tr712 virion. Producer H9 (np) cells had been weakly infected (less than 20% infection level). A. Western blot: the top part of the filter has been probed with gp120 antibodies, the middle part with gp41 mAb, Chessie 8 against the Env C-terminal tail (truncated in HIV-Env-Tr712) and the bottom part with p24 mAb. B. Average gp120 incorporation into HIV-Env-Tr712 virions in comparison to HIV-Wt (from 3 independent experiments: individual values 37%, 118%, 83%).
Mentions: Additionally, it is likely that differences in infection levels of producer cells also account for the fact that, in contrast to our earlier report [33], others had previously reported reduction in the cell-free infectivity of HIV-Env-Tr712 virions produced in non-permissive cells [19,21]. In these latter studies, reduced infectivity had been reported to correlate with a defect in Env-Tr712 incorporation [19,21]. Thus, in order to study this here, HIV-Wt and HIV-Env-Tr712 virions were produced in weakly infected H9 (np) cells (less than 20% infected) and their protein content analysed by Western blot. Three independent experiments were evaluated by quantifying the amount of incorporated gp120 relative to viral p24 content for each virus preparation. As seen in Fig. 3A, gp120 incorporation into HIV-Env-Tr712 virions can clearly be seen and, as shown in Fig. 3B, the amount is, on average 79% of that in HIV-Wt. The reason for this discrepancy between these Env incorporation results and those previously published is presently not known. At any rate, this modest reduction in Env incorporation appears unlikely to account for the strongly reduced infectivity of cell-free HIV-Env-Tr712 virions.

Bottom Line: We further observed that in permissive cells, which supported both routes of mutant virus transmission, viral gene expression levels, Gag processing and particle release were inherently higher than in non-permissive cells, a factor which may be significantly contributing to their permissivity phenotype.Additionally, and correlating with viral transfer efficiencies in these cell types, HIV-Gag accumulation at the virological synapse (VS) was reduced to background levels in the absence of the Env-CT in conjugates of non-permissive cells but not in permissive cells.The requirement of the Env-CT for these related processes is abrogated in permissive cells, which exhibit higher HIV gene expression levels.

View Article: PubMed Central - HTML - PubMed

Affiliation: Forschungsschwerpunkt Infektion und Krebs, F020, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.

ABSTRACT

Background: Mutant HIV (HIV-Env-Tr712) lacking the cytoplasmic tail of the viral glycoprotein (Env-CT) exhibits a cell-type specific replication phenotype such that replicative spread occurs in some T-cell lines (referred to as permissive cells) but fails to do so in most T-cell lines or in PBMCs (referred to as non-permissive cells). We aim to gain insight on the underlying requirement for the Env-CT for viral spread in non-permissive cells.

Results: We established that in comparison to HIV-Wt, both cell-free and cell-to-cell transmission of mutant HIV-Env-Tr712 from non-permissive cells were severely impaired under naturally low infection conditions. This requirement for Env-CT could be largely overcome by using saturating amounts of virus for infection. We further observed that in permissive cells, which supported both routes of mutant virus transmission, viral gene expression levels, Gag processing and particle release were inherently higher than in non-permissive cells, a factor which may be significantly contributing to their permissivity phenotype. Additionally, and correlating with viral transfer efficiencies in these cell types, HIV-Gag accumulation at the virological synapse (VS) was reduced to background levels in the absence of the Env-CT in conjugates of non-permissive cells but not in permissive cells.

Conclusions: During natural infection conditions, the HIV-Env-CT is critically required for viral transmission in cultures of non-permissive cells by both cell-free and cell-to-cell routes and is instrumental for Gag accumulation to the VS. The requirement of the Env-CT for these related processes is abrogated in permissive cells, which exhibit higher HIV gene expression levels.

Show MeSH
Related in: MedlinePlus