PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells

Maria Lopez-Huertas, Jasmine Li, Anjum ZAFAR, Sara Rodriguez-Mora, Carlota Garcia-Dominguez, Elena Mateos, Jose Alcami, Sudha RAO, Mayte Coiras

Research output: Contribution to journalArticle

4 Citations (Scopus)
3 Downloads (Pure)

Abstract

PKCθ is essential for the activation of CD4 + T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4 + T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T 538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4 + T cells and could be used as a therapeutic target.

Original languageEnglish
Article number69
Pages (from-to)1-14
Number of pages14
JournalFrontiers in Immunology
Volume7
Issue numberFEB
DOIs
Publication statusPublished - 2016

Fingerprint

HIV-1
HIV Long Terminal Repeat
T-Lymphocytes
Jurkat Cells
Doxycycline
Transcriptional Activation
Immunological Synapses
Messenger RNA
MAP Kinase Signaling System
RNA Interference
Metabolic Networks and Pathways
Cytosol
Transcription Factors
Phosphotransferases
Up-Regulation
Phosphorylation
Cell Membrane
RNA
Therapeutics

Cite this

Lopez-Huertas, M., Li, J., ZAFAR, A., Rodriguez-Mora, S., Garcia-Dominguez, C., Mateos, E., ... Coiras, M. (2016). PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells. Frontiers in Immunology, 7(FEB), 1-14. [69]. https://doi.org/10.3389/fimmu.2016.00069
Lopez-Huertas, Maria ; Li, Jasmine ; ZAFAR, Anjum ; Rodriguez-Mora, Sara ; Garcia-Dominguez, Carlota ; Mateos, Elena ; Alcami, Jose ; RAO, Sudha ; Coiras, Mayte. / PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells. In: Frontiers in Immunology. 2016 ; Vol. 7, No. FEB. pp. 1-14.
@article{578034c10a3849dc90f8305156f3d5b9,
title = "PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells",
abstract = "PKCθ is essential for the activation of CD4 + T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4 + T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T 538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4 + T cells and could be used as a therapeutic target.",
keywords = "CD4+ T cell activation, HIV-1 Tat regulator, NF-kappa B, Nuclear colocalization, PKC theta and HIV-1 Tat interaction, PKC theta mRNA interference, Protein kinase C theta, Ras/Raf/MEK/Erk pathway",
author = "Maria Lopez-Huertas and Jasmine Li and Anjum ZAFAR and Sara Rodriguez-Mora and Carlota Garcia-Dominguez and Elena Mateos and Jose Alcami and Sudha RAO and Mayte Coiras",
year = "2016",
doi = "10.3389/fimmu.2016.00069",
language = "English",
volume = "7",
pages = "1--14",
journal = "Frontiers in Immunology",
issn = "1664-3224",
publisher = "Frontiers Media S.A.",
number = "FEB",

}

Lopez-Huertas, M, Li, J, ZAFAR, A, Rodriguez-Mora, S, Garcia-Dominguez, C, Mateos, E, Alcami, J, RAO, S & Coiras, M 2016, 'PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells', Frontiers in Immunology, vol. 7, no. FEB, 69, pp. 1-14. https://doi.org/10.3389/fimmu.2016.00069

PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells. / Lopez-Huertas, Maria; Li, Jasmine; ZAFAR, Anjum; Rodriguez-Mora, Sara; Garcia-Dominguez, Carlota; Mateos, Elena; Alcami, Jose; RAO, Sudha; Coiras, Mayte.

In: Frontiers in Immunology, Vol. 7, No. FEB, 69, 2016, p. 1-14.

Research output: Contribution to journalArticle

TY - JOUR

T1 - PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells

AU - Lopez-Huertas, Maria

AU - Li, Jasmine

AU - ZAFAR, Anjum

AU - Rodriguez-Mora, Sara

AU - Garcia-Dominguez, Carlota

AU - Mateos, Elena

AU - Alcami, Jose

AU - RAO, Sudha

AU - Coiras, Mayte

PY - 2016

Y1 - 2016

N2 - PKCθ is essential for the activation of CD4 + T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4 + T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T 538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4 + T cells and could be used as a therapeutic target.

AB - PKCθ is essential for the activation of CD4 + T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4 + T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T 538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4 + T cells and could be used as a therapeutic target.

KW - CD4+ T cell activation

KW - HIV-1 Tat regulator

KW - NF-kappa B

KW - Nuclear colocalization

KW - PKC theta and HIV-1 Tat interaction

KW - PKC theta mRNA interference

KW - Protein kinase C theta

KW - Ras/Raf/MEK/Erk pathway

UR - http://www.scopus.com/inward/record.url?scp=84962529747&partnerID=8YFLogxK

U2 - 10.3389/fimmu.2016.00069

DO - 10.3389/fimmu.2016.00069

M3 - Article

VL - 7

SP - 1

EP - 14

JO - Frontiers in Immunology

JF - Frontiers in Immunology

SN - 1664-3224

IS - FEB

M1 - 69

ER -

Lopez-Huertas M, Li J, ZAFAR A, Rodriguez-Mora S, Garcia-Dominguez C, Mateos E et al. PKC-omerga and HIV-1 transcriptional regulator Tat co-exist at the LTR promoter in CD4+ T cells. Frontiers in Immunology. 2016;7(FEB):1-14. 69. https://doi.org/10.3389/fimmu.2016.00069