Limits...
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: 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.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.

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

ImageStream analysis of Foxo1 and NF-κB in HIV-1 infected cells.HIV-1 GFP-reporter virus-infected resting CD4+ T cells were stained for CD45RA, intracellular Foxo1 or NF-κB and for DNA (DAPI) 7 days after infection, then analyzed by ImageStream. One representative experiment of two is shown. A. Total Foxo1 expression in naïve and memory GFP+ and GFP- cells. MFI = Mean Fluorescence Intensity. B. Nuclear localization was analyzed in IDEAS software by comparison of Foxo1 distribution to nuclear DAPI staining using the Similarity Score (see materials and methods). C. Representative ImageStream pictures of individual cells stained for Foxo1. D. Total NF-κB expression in naïve and memory GFP+ and GFP- cells. E. NF-κB nuclear Similarity Scores. F. Representative ImageStream pictures of individual cells stained for NF-κB.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4199762&req=5

pone-0110719-g004: ImageStream analysis of Foxo1 and NF-κB in HIV-1 infected cells.HIV-1 GFP-reporter virus-infected resting CD4+ T cells were stained for CD45RA, intracellular Foxo1 or NF-κB and for DNA (DAPI) 7 days after infection, then analyzed by ImageStream. One representative experiment of two is shown. A. Total Foxo1 expression in naïve and memory GFP+ and GFP- cells. MFI = Mean Fluorescence Intensity. B. Nuclear localization was analyzed in IDEAS software by comparison of Foxo1 distribution to nuclear DAPI staining using the Similarity Score (see materials and methods). C. Representative ImageStream pictures of individual cells stained for Foxo1. D. Total NF-κB expression in naïve and memory GFP+ and GFP- cells. E. NF-κB nuclear Similarity Scores. F. Representative ImageStream pictures of individual cells stained for NF-κB.

Mentions: Freed from DNA binding, inactivated Foxo1 relocalizes to the cytoplasm; thus cytoplasmic translocation is a direct indication of Foxo1 inactivation. We next examined Foxo1 localization by ImageStream analysis, a technique that combines flow cytometry with cell imaging to provide spatial and intensity information within individual cells (Figure 4). We first observed that total Foxo1 levels were not altered in the productively infected naïve CD4 T cells, and slightly increased in memory cells (Figure 4A). When nuclear and cytoplasmic expression were analyzed, translocation of Foxo1 from nucleus to the (very thin) cytoplasm, was observed in both the naïve and memory resting T cells (Figure 4B). This translocation was clearly evident in images of individual cells (Figure 4C). By contrast with Foxo1, NF-κB remained almost completely cytoplasmic (Figure 4D–F). However, similar to Foxo1, NF-κB total levels were increased in the GFP+ memory cells (Figure 4D), though no nuclear translocation was observed. GFP+ cells tended to be more elongated and irregular in shape, resembling polarized T cells, a further indication of partial activation (see the middle cells in memory GFP+ cells stained for Foxo1 in Figure 4C).


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)

ImageStream analysis of Foxo1 and NF-κB in HIV-1 infected cells.HIV-1 GFP-reporter virus-infected resting CD4+ T cells were stained for CD45RA, intracellular Foxo1 or NF-κB and for DNA (DAPI) 7 days after infection, then analyzed by ImageStream. One representative experiment of two is shown. A. Total Foxo1 expression in naïve and memory GFP+ and GFP- cells. MFI = Mean Fluorescence Intensity. B. Nuclear localization was analyzed in IDEAS software by comparison of Foxo1 distribution to nuclear DAPI staining using the Similarity Score (see materials and methods). C. Representative ImageStream pictures of individual cells stained for Foxo1. D. Total NF-κB expression in naïve and memory GFP+ and GFP- cells. E. NF-κB nuclear Similarity Scores. F. Representative ImageStream pictures of individual cells stained for NF-κB.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4199762&req=5

pone-0110719-g004: ImageStream analysis of Foxo1 and NF-κB in HIV-1 infected cells.HIV-1 GFP-reporter virus-infected resting CD4+ T cells were stained for CD45RA, intracellular Foxo1 or NF-κB and for DNA (DAPI) 7 days after infection, then analyzed by ImageStream. One representative experiment of two is shown. A. Total Foxo1 expression in naïve and memory GFP+ and GFP- cells. MFI = Mean Fluorescence Intensity. B. Nuclear localization was analyzed in IDEAS software by comparison of Foxo1 distribution to nuclear DAPI staining using the Similarity Score (see materials and methods). C. Representative ImageStream pictures of individual cells stained for Foxo1. D. Total NF-κB expression in naïve and memory GFP+ and GFP- cells. E. NF-κB nuclear Similarity Scores. F. Representative ImageStream pictures of individual cells stained for NF-κB.
Mentions: Freed from DNA binding, inactivated Foxo1 relocalizes to the cytoplasm; thus cytoplasmic translocation is a direct indication of Foxo1 inactivation. We next examined Foxo1 localization by ImageStream analysis, a technique that combines flow cytometry with cell imaging to provide spatial and intensity information within individual cells (Figure 4). We first observed that total Foxo1 levels were not altered in the productively infected naïve CD4 T cells, and slightly increased in memory cells (Figure 4A). When nuclear and cytoplasmic expression were analyzed, translocation of Foxo1 from nucleus to the (very thin) cytoplasm, was observed in both the naïve and memory resting T cells (Figure 4B). This translocation was clearly evident in images of individual cells (Figure 4C). By contrast with Foxo1, NF-κB remained almost completely cytoplasmic (Figure 4D–F). However, similar to Foxo1, NF-κB total levels were increased in the GFP+ memory cells (Figure 4D), though no nuclear translocation was observed. GFP+ cells tended to be more elongated and irregular in shape, resembling polarized T cells, a further indication of partial activation (see the middle cells in memory GFP+ cells stained for Foxo1 in Figure 4C).

Bottom Line: 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.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.

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