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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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Replicative spread of HIV-Wt and HIV-Env-Tr712 in different T-cells. A. Schematic depiction of the Env proteins from HIV-Wt and HIV-Env-Tr712 B. Replication kinetics of HIV-Wt (circles) and HIV-Env-Tr712 (triangles) in MT-4 cells (p) and H9 cells (np) below. Cells were infected with equal amounts of virus (equivalent to 100 ng p24 per 106 cells) and washed at 5 h p.i. Newly produced virions released into the culture supernatants at the times indicated were quantified by CA-ELISA. Note that at 3-4 d post infection, MT-4 cells were infected to 100% with both viruses (as established by indirect immunofluorescence) and succumbed to HIV induced cytotoxicity. Infection of H9 cells with HIV-Wt reached 100% at 4-5 d post-infection whereas infection with HIV-Env-Tr712 virions (produced in permissive 293T cells) resulted in initial infection of only a low percentage (< 5%) of cells which subsequently vanished from the culture. The cut-off of the assay lies at 0.01 ng/ml.
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Figure 1: Replicative spread of HIV-Wt and HIV-Env-Tr712 in different T-cells. A. Schematic depiction of the Env proteins from HIV-Wt and HIV-Env-Tr712 B. Replication kinetics of HIV-Wt (circles) and HIV-Env-Tr712 (triangles) in MT-4 cells (p) and H9 cells (np) below. Cells were infected with equal amounts of virus (equivalent to 100 ng p24 per 106 cells) and washed at 5 h p.i. Newly produced virions released into the culture supernatants at the times indicated were quantified by CA-ELISA. Note that at 3-4 d post infection, MT-4 cells were infected to 100% with both viruses (as established by indirect immunofluorescence) and succumbed to HIV induced cytotoxicity. Infection of H9 cells with HIV-Wt reached 100% at 4-5 d post-infection whereas infection with HIV-Env-Tr712 virions (produced in permissive 293T cells) resulted in initial infection of only a low percentage (< 5%) of cells which subsequently vanished from the culture. The cut-off of the assay lies at 0.01 ng/ml.

Mentions: Fig 1A schematically depicts the Env proteins of HIV-Wt and HIV-Env-Tr712. As detailed in the Introduction and shown in Fig. 1B, HIV-Env-Tr712 virions exhibit a cell-type specific defect such that replicative spread occurs in some T-cell lines (here MT-4 cells, termed permissive (p) cells) but fails to occur in the majority of T-cell lines (here H9 cells, referred to as non-permissive (np) cells). In this report, our emphasis was to analyse the possible impact of Env-CT truncation on cell-to-cell viral transmission in both H9 (np) and MT-4 (p) cells. Infected donor cells were generated by infection with VSV-G pseudotyped Wt or mutant viruses and, in the course of our studies, we observed that their initial infection level markedly influenced experimental outcome. Thus, in this report, we describe cell-to-cell transmission experiments employing highly or weakly infected donor cells and have also analysed cell-free viral infectivities in the context of both of these scenarios.


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)

Replicative spread of HIV-Wt and HIV-Env-Tr712 in different T-cells. A. Schematic depiction of the Env proteins from HIV-Wt and HIV-Env-Tr712 B. Replication kinetics of HIV-Wt (circles) and HIV-Env-Tr712 (triangles) in MT-4 cells (p) and H9 cells (np) below. Cells were infected with equal amounts of virus (equivalent to 100 ng p24 per 106 cells) and washed at 5 h p.i. Newly produced virions released into the culture supernatants at the times indicated were quantified by CA-ELISA. Note that at 3-4 d post infection, MT-4 cells were infected to 100% with both viruses (as established by indirect immunofluorescence) and succumbed to HIV induced cytotoxicity. Infection of H9 cells with HIV-Wt reached 100% at 4-5 d post-infection whereas infection with HIV-Env-Tr712 virions (produced in permissive 293T cells) resulted in initial infection of only a low percentage (< 5%) of cells which subsequently vanished from the culture. The cut-off of the assay lies at 0.01 ng/ml.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 1: Replicative spread of HIV-Wt and HIV-Env-Tr712 in different T-cells. A. Schematic depiction of the Env proteins from HIV-Wt and HIV-Env-Tr712 B. Replication kinetics of HIV-Wt (circles) and HIV-Env-Tr712 (triangles) in MT-4 cells (p) and H9 cells (np) below. Cells were infected with equal amounts of virus (equivalent to 100 ng p24 per 106 cells) and washed at 5 h p.i. Newly produced virions released into the culture supernatants at the times indicated were quantified by CA-ELISA. Note that at 3-4 d post infection, MT-4 cells were infected to 100% with both viruses (as established by indirect immunofluorescence) and succumbed to HIV induced cytotoxicity. Infection of H9 cells with HIV-Wt reached 100% at 4-5 d post-infection whereas infection with HIV-Env-Tr712 virions (produced in permissive 293T cells) resulted in initial infection of only a low percentage (< 5%) of cells which subsequently vanished from the culture. The cut-off of the assay lies at 0.01 ng/ml.
Mentions: Fig 1A schematically depicts the Env proteins of HIV-Wt and HIV-Env-Tr712. As detailed in the Introduction and shown in Fig. 1B, HIV-Env-Tr712 virions exhibit a cell-type specific defect such that replicative spread occurs in some T-cell lines (here MT-4 cells, termed permissive (p) cells) but fails to occur in the majority of T-cell lines (here H9 cells, referred to as non-permissive (np) cells). In this report, our emphasis was to analyse the possible impact of Env-CT truncation on cell-to-cell viral transmission in both H9 (np) and MT-4 (p) cells. Infected donor cells were generated by infection with VSV-G pseudotyped Wt or mutant viruses and, in the course of our studies, we observed that their initial infection level markedly influenced experimental outcome. Thus, in this report, we describe cell-to-cell transmission experiments employing highly or weakly infected donor cells and have also analysed cell-free viral infectivities in the context of both of these scenarios.

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