Cargando…
PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells
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 demon...
Autores principales: | , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2016
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4770193/ https://www.ncbi.nlm.nih.gov/pubmed/26973648 http://dx.doi.org/10.3389/fimmu.2016.00069 |
_version_ | 1782418215648886784 |
---|---|
author | López-Huertas, María Rosa Li, Jasmine Zafar, Anjum Rodríguez-Mora, Sara García-Domínguez, Carlota Mateos, Elena Alcamí, José Rao, Sudha Coiras, Mayte |
author_facet | López-Huertas, María Rosa Li, Jasmine Zafar, Anjum Rodríguez-Mora, Sara García-Domínguez, Carlota Mateos, Elena Alcamí, José Rao, Sudha Coiras, Mayte |
author_sort | López-Huertas, María Rosa |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-4770193 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-47701932016-03-11 PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells López-Huertas, María Rosa Li, Jasmine Zafar, Anjum Rodríguez-Mora, Sara García-Domínguez, Carlota Mateos, Elena Alcamí, José Rao, Sudha Coiras, Mayte Front Immunol Immunology 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. Frontiers Media S.A. 2016-02-29 /pmc/articles/PMC4770193/ /pubmed/26973648 http://dx.doi.org/10.3389/fimmu.2016.00069 Text en Copyright © 2016 López-Huertas, Li, Zafar, Rodríguez-Mora, García-Domínguez, Mateos, Alcamí, Rao and Coiras. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Immunology López-Huertas, María Rosa Li, Jasmine Zafar, Anjum Rodríguez-Mora, Sara García-Domínguez, Carlota Mateos, Elena Alcamí, José Rao, Sudha Coiras, Mayte PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title | PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title_full | PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title_fullStr | PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title_full_unstemmed | PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title_short | PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4(+) T Cells |
title_sort | pkcθ and hiv-1 transcriptional regulator tat co-exist at the ltr promoter in cd4(+) t cells |
topic | Immunology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4770193/ https://www.ncbi.nlm.nih.gov/pubmed/26973648 http://dx.doi.org/10.3389/fimmu.2016.00069 |
work_keys_str_mv | AT lopezhuertasmariarosa pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT lijasmine pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT zafaranjum pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT rodriguezmorasara pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT garciadominguezcarlota pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT mateoselena pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT alcamijose pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT raosudha pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells AT coirasmayte pkcthandhiv1transcriptionalregulatortatcoexistattheltrpromoterincd4tcells |