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Suppression of Foxo1 activity and down-modulation of CD62L (L-selectin) in HIV-1 infected resting CD4 T cells.

Trinité B, Chan CN, Lee CS, Mahajan S, Luo Y, Muesing MA, Folkvord JM, Pham M, Connick E, Levy DN - PLoS ONE (2014)

Bottom Line: CD62L gene transcription is maintained in resting T cells by Foxo1 and KLF2, transcription factors that maintain T cell quiescence and which regulate additional cellular processes including survival, migration, and differentiation.Partial T cell activation was further evident as an increase in CD69 expression.Several other Foxo1- and KLF2-regulated mRNA were increased or decreased in productively infected CD4 T cells, including IL-7rα, Myc, CCR5, Fam65b, S1P1 (EDG1), CD52, Cyclin D2 and p21CIP1, indicating a profound reprogramming of these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Basic Science, New York University College of Dentistry, New York, New York, United States of America.

ABSTRACT
HIV-1 hijacks and disrupts many processes in the cells it infects in order to suppress antiviral immunity and to facilitate its replication. Resting CD4 T cells are important early targets of HIV-1 infection in which HIV-1 must overcome intrinsic barriers to viral replication. Although resting CD4 T cells are refractory to infection in vitro, local environmental factors within lymphoid and mucosal tissues such as cytokines facilitate viral replication while maintaining the resting state. These factors can be utilized in vitro to study HIV-1 replication in resting CD4 T cells. In vivo, the migration of resting naïve and central memory T cells into lymphoid tissues is dependent upon expression of CD62L (L-selectin), a receptor that is subsequently down-modulated following T cell activation. CD62L gene transcription is maintained in resting T cells by Foxo1 and KLF2, transcription factors that maintain T cell quiescence and which regulate additional cellular processes including survival, migration, and differentiation. Here we report that HIV-1 down-modulates CD62L in productively infected naïve and memory resting CD4 T cells while suppressing Foxo1 activity and the expression of KLF2 mRNA. Partial T cell activation was further evident as an increase in CD69 expression. Several other Foxo1- and KLF2-regulated mRNA were increased or decreased in productively infected CD4 T cells, including IL-7rα, Myc, CCR5, Fam65b, S1P1 (EDG1), CD52, Cyclin D2 and p21CIP1, indicating a profound reprogramming of these cells. The Foxo1 inhibitor AS1842856 accelerated de novo viral gene expression and the sequella of infection, supporting the notion that HIV-1 suppression of Foxo1 activity may be a strategy to promote replication in resting CD4 T cells. As Foxo1 is an investigative cancer therapy target, the development of Foxo1 interventions may assist the quest to specifically suppress or activate HIV-1 replication in vivo.

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Down-modulation of CD62L and upregulation of CD69 and on productively infected naïve and memory resting CD4 T cells.A. Resting IL-7 treated CD4+ T cells were infected with the GFP reporter virus NLENG1, restricted to a single round of infection by treatment with the protease inhibitor indinavir (2 µM). Flow analysis was carried out on day 5 post infection on cells with and without activation by αCD3/CD28 beads on day 3 p.i. Grey: isotype-matched IgG control antibody staining. Blue: GFP-negative cells. Green: GFP+ cells. Data are representative of 4 independent experiments. B. Fold change in CD62L and CD69 expression on naïve and memory CD4+ resting T cells in A. With on exception (*) all values were statistically different (p<0.01) from the negative control by a T(x) population comparison test [134]. C. Failure of GFP+ cells proliferate demonstrates that CD62L-low cells are not produced by preferential expansion of an existing CD62L-low population. Experiment shown is representative of >5 similar experiments. D. IL-4 supports HIV-1-induced down-modulation of CD62L on both naïve and memory resting CD4+ T cells. Experiment shown is representative of >3 similar experiments. E. CD62L down-modulation on HIV-1 infected naïve tonsil T cells 7 days after infection. Similar results were observed on CD45R0+cells (not shown). Experiment shown is representative of 2 similar experiments.
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pone-0110719-g001: Down-modulation of CD62L and upregulation of CD69 and on productively infected naïve and memory resting CD4 T cells.A. Resting IL-7 treated CD4+ T cells were infected with the GFP reporter virus NLENG1, restricted to a single round of infection by treatment with the protease inhibitor indinavir (2 µM). Flow analysis was carried out on day 5 post infection on cells with and without activation by αCD3/CD28 beads on day 3 p.i. Grey: isotype-matched IgG control antibody staining. Blue: GFP-negative cells. Green: GFP+ cells. Data are representative of 4 independent experiments. B. Fold change in CD62L and CD69 expression on naïve and memory CD4+ resting T cells in A. With on exception (*) all values were statistically different (p<0.01) from the negative control by a T(x) population comparison test [134]. C. Failure of GFP+ cells proliferate demonstrates that CD62L-low cells are not produced by preferential expansion of an existing CD62L-low population. Experiment shown is representative of >5 similar experiments. D. IL-4 supports HIV-1-induced down-modulation of CD62L on both naïve and memory resting CD4+ T cells. Experiment shown is representative of >3 similar experiments. E. CD62L down-modulation on HIV-1 infected naïve tonsil T cells 7 days after infection. Similar results were observed on CD45R0+cells (not shown). Experiment shown is representative of 2 similar experiments.

Mentions: We next examined the influence of HIV-1 infection on expression of CD4, CD69 and CD62L on resting CD4+ T cells (Figure 1). We activated a subset of cells with anti-CD3/CD28 beads 2 days after infection in order to compare receptor expression on infected resting vs. activated cells. CD4 was efficiently down-modulated by the virus, as expected, whether the cells were activated or not after infection (Figure 1A). Interestingly, the early activation marker CD69 was modestly upregulated in both memory (CD4+CD45RA+) and naïve (CD4+CD45RA−) resting GFP+ cells, (Figure 1B). There was variability in the effect of infection on both receptors among cell donors, but with a single exception, all infections produced significant CD62L down modulation and CD69 upregulation. Late activation markers such as CD25, CD38 and HLA Class II were unaffected (not shown). Mock infected cells remained uniformly CD69-negative (not shown), thus HIV-1 is actively inducing CD69 expression.


Suppression of Foxo1 activity and down-modulation of CD62L (L-selectin) in HIV-1 infected resting CD4 T cells.

Trinité B, Chan CN, Lee CS, Mahajan S, Luo Y, Muesing MA, Folkvord JM, Pham M, Connick E, Levy DN - PLoS ONE (2014)

Down-modulation of CD62L and upregulation of CD69 and on productively infected naïve and memory resting CD4 T cells.A. Resting IL-7 treated CD4+ T cells were infected with the GFP reporter virus NLENG1, restricted to a single round of infection by treatment with the protease inhibitor indinavir (2 µM). Flow analysis was carried out on day 5 post infection on cells with and without activation by αCD3/CD28 beads on day 3 p.i. Grey: isotype-matched IgG control antibody staining. Blue: GFP-negative cells. Green: GFP+ cells. Data are representative of 4 independent experiments. B. Fold change in CD62L and CD69 expression on naïve and memory CD4+ resting T cells in A. With on exception (*) all values were statistically different (p<0.01) from the negative control by a T(x) population comparison test [134]. C. Failure of GFP+ cells proliferate demonstrates that CD62L-low cells are not produced by preferential expansion of an existing CD62L-low population. Experiment shown is representative of >5 similar experiments. D. IL-4 supports HIV-1-induced down-modulation of CD62L on both naïve and memory resting CD4+ T cells. Experiment shown is representative of >3 similar experiments. E. CD62L down-modulation on HIV-1 infected naïve tonsil T cells 7 days after infection. Similar results were observed on CD45R0+cells (not shown). Experiment shown is representative of 2 similar experiments.
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Related In: Results  -  Collection

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pone-0110719-g001: Down-modulation of CD62L and upregulation of CD69 and on productively infected naïve and memory resting CD4 T cells.A. Resting IL-7 treated CD4+ T cells were infected with the GFP reporter virus NLENG1, restricted to a single round of infection by treatment with the protease inhibitor indinavir (2 µM). Flow analysis was carried out on day 5 post infection on cells with and without activation by αCD3/CD28 beads on day 3 p.i. Grey: isotype-matched IgG control antibody staining. Blue: GFP-negative cells. Green: GFP+ cells. Data are representative of 4 independent experiments. B. Fold change in CD62L and CD69 expression on naïve and memory CD4+ resting T cells in A. With on exception (*) all values were statistically different (p<0.01) from the negative control by a T(x) population comparison test [134]. C. Failure of GFP+ cells proliferate demonstrates that CD62L-low cells are not produced by preferential expansion of an existing CD62L-low population. Experiment shown is representative of >5 similar experiments. D. IL-4 supports HIV-1-induced down-modulation of CD62L on both naïve and memory resting CD4+ T cells. Experiment shown is representative of >3 similar experiments. E. CD62L down-modulation on HIV-1 infected naïve tonsil T cells 7 days after infection. Similar results were observed on CD45R0+cells (not shown). Experiment shown is representative of 2 similar experiments.
Mentions: We next examined the influence of HIV-1 infection on expression of CD4, CD69 and CD62L on resting CD4+ T cells (Figure 1). We activated a subset of cells with anti-CD3/CD28 beads 2 days after infection in order to compare receptor expression on infected resting vs. activated cells. CD4 was efficiently down-modulated by the virus, as expected, whether the cells were activated or not after infection (Figure 1A). Interestingly, the early activation marker CD69 was modestly upregulated in both memory (CD4+CD45RA+) and naïve (CD4+CD45RA−) resting GFP+ cells, (Figure 1B). There was variability in the effect of infection on both receptors among cell donors, but with a single exception, all infections produced significant CD62L down modulation and CD69 upregulation. Late activation markers such as CD25, CD38 and HLA Class II were unaffected (not shown). Mock infected cells remained uniformly CD69-negative (not shown), thus HIV-1 is actively inducing CD69 expression.

Bottom Line: CD62L gene transcription is maintained in resting T cells by Foxo1 and KLF2, transcription factors that maintain T cell quiescence and which regulate additional cellular processes including survival, migration, and differentiation.Partial T cell activation was further evident as an increase in CD69 expression.Several other Foxo1- and KLF2-regulated mRNA were increased or decreased in productively infected CD4 T cells, including IL-7rα, Myc, CCR5, Fam65b, S1P1 (EDG1), CD52, Cyclin D2 and p21CIP1, indicating a profound reprogramming of these cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Basic Science, New York University College of Dentistry, New York, New York, United States of America.

ABSTRACT
HIV-1 hijacks and disrupts many processes in the cells it infects in order to suppress antiviral immunity and to facilitate its replication. Resting CD4 T cells are important early targets of HIV-1 infection in which HIV-1 must overcome intrinsic barriers to viral replication. Although resting CD4 T cells are refractory to infection in vitro, local environmental factors within lymphoid and mucosal tissues such as cytokines facilitate viral replication while maintaining the resting state. These factors can be utilized in vitro to study HIV-1 replication in resting CD4 T cells. In vivo, the migration of resting naïve and central memory T cells into lymphoid tissues is dependent upon expression of CD62L (L-selectin), a receptor that is subsequently down-modulated following T cell activation. CD62L gene transcription is maintained in resting T cells by Foxo1 and KLF2, transcription factors that maintain T cell quiescence and which regulate additional cellular processes including survival, migration, and differentiation. Here we report that HIV-1 down-modulates CD62L in productively infected naïve and memory resting CD4 T cells while suppressing Foxo1 activity and the expression of KLF2 mRNA. Partial T cell activation was further evident as an increase in CD69 expression. Several other Foxo1- and KLF2-regulated mRNA were increased or decreased in productively infected CD4 T cells, including IL-7rα, Myc, CCR5, Fam65b, S1P1 (EDG1), CD52, Cyclin D2 and p21CIP1, indicating a profound reprogramming of these cells. The Foxo1 inhibitor AS1842856 accelerated de novo viral gene expression and the sequella of infection, supporting the notion that HIV-1 suppression of Foxo1 activity may be a strategy to promote replication in resting CD4 T cells. As Foxo1 is an investigative cancer therapy target, the development of Foxo1 interventions may assist the quest to specifically suppress or activate HIV-1 replication in vivo.

Show MeSH
Related in: MedlinePlus