Inhibition of Human Immunodeficiency Virus Type 1 Replication in Latently Infected Cells by a Novel IκB Kinase Inhibitor

Victoriano, Ann Florence B.; Asamitsu, Kaori; Hibi, Yurina; Imai, Kenichi; Barzaga, Nina G.; Okamoto, Takashi

doi:10.1128/aac.50.2.547-555.2006

Cited by 34 publications

(29 citation statements)

References 76 publications

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“…Chromatin Immunoprecipitation (ChIP) Assay-ChIP assays were performed according to the provider's protocol (Upstate Biotechnology) with some modifications as previously described (18,20,40,42). In brief, chromatin from cross-linked cells was sheared by sonication 13 times for 10 s at one-third of the maximum power of a Microson XL sonicator (Wakenyaku, Co., Ltd., Kyoto, Japan) with 20 s cooling on ice between each pulse and incubated with specific antibody followed by incubation with protein A-agarose beads saturated with salmon sperm DNA.…”

Section: Methodsmentioning

confidence: 99%

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Involvement of Histone H3 Lysine 9 (H3K9) Methyltransferase G9a in the Maintenance of HIV-1 Latency and Its Reactivation by BIX01294

Imai

Togami

Okamoto

2010

Journal of Biological Chemistry

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222

229

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Elucidating the mechanism of human immunodeficiency virus, type 1 (HIV-1) provirus transcriptional silencing in latently infected cells is crucial for understanding the pathophysiological process of HIV-1 infection. It is well established that hypoacetylation of histone proteins by histone deacetylases is involved in the maintenance of HIV-1 latency by repressing viral transcription. Although histone methylation is involved in the organization of chromatin domains and plays a central epigenetic role in gene expression, the role of histone methylation in the maintenance of HIV-1 latency has not been clarified. Here we present evidence that histone H3 Lys 9 (H3K9) methyltransferase G9a is responsible for transcriptional repression of HIV-1 by promoting repressive dimethylation at H3K9 and for the maintenance of viral latency. We observed that G9a significantly inhibited basal, as well as, the induced HIV-1 gene expression by tumor necrosis factor-␣ or Tat. Mutant G9a, however, lacking the SET domain responsible for the catalytic activity of histone methyltransferase, did not show such an effect. When G9a expression was knocked down by small interfering RNA, HIV-1 replication was augmented from cells transiently transfected with a full-length HIV-1 clone. Moreover, a specific inhibitor of G9a, BIX01294, could reactivate expression of HIV-1 from latently infected cells such as ACH-2 and OM10.1. Furthermore, chromatin immunoprecipitation assays revealed the presence of G9a and H3K9 dimethylation on nucleosome histones in the vicinity of the HIV-1 long terminal repeat promoter. These results suggest that G9a is responsible for the transcriptional quiescence of latent HIV-1 provirus and provide a molecular basis for understanding the mechanism by which HIV-1 latency is maintained.

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Section: Methodsmentioning

confidence: 99%

“…Measurement of Viral p24 Antigen-The p24 antigen level in the cell culture supernatant was measured by p24 antigen capture ELISA using a RETRO-TEK HIV-1 p24 Antigen ELISA kit (Zepto Metrix Corp., Buffalo, NY) as described previously (18,20,42). All experiments were performed in triplicates and the data presented as mean Ϯ S.D.…”

Section: Methodsmentioning

confidence: 99%

Involvement of Histone H3 Lysine 9 (H3K9) Methyltransferase G9a in the Maintenance of HIV-1 Latency and Its Reactivation by BIX01294

Imai

Togami

Okamoto

2010

Journal of Biological Chemistry

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222

229

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“…The p24 antigen level in the cell culture supernatant was measured by a p24 antigen capture ELISA assay using a commercial kit (RETRO-TEK HIV-1 p24 Antigen ELISA kit; Zepto Metrix , Buffalo, NY, USA) [28,32].…”

Section: Measurement Of Viral P24 Antigenmentioning

confidence: 99%

Reactivation of latent HIV-1 by a wide variety of butyric acid-producing bacteria

Imai

Yamada

Tamura

et al. 2012

Cell. Mol. Life Sci.

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Latently infected cells harbor human immunodeficiency virus type 1 (HIV-1) proviral DNA copies integrated in heterochromatin, allowing persistence of transcriptionally silent proviruses. It is widely accepted that hypoacetylation of histone proteins by histone deacetylases (HDACs) is involved in maintaining the HIV-1 latency by repressing viral transcription. HIV-1 replication can be induced from latently infected cells by environmental factors, such as inflammation and co-infection with other microbes. It is known that a bacterial metabolite butyric acid inhibits catalytic action of HDAC and induces transcription of silenced genes including HIV-1 provirus. There are a number of such bacteria in gut, vaginal, and oral cavities that produce butyric acid during their anaerobic glycolysis. Since these organs are known to be the major site of HIV-1 transmission and its replication, we explored a possibility that explosive viral replication in these organs could be ascribable to butyric acid produced from anaerobic resident bacteria. In this study, we demonstrate that the culture supernatant of various bacteria producing butyric acid could greatly reactivate the latently-infected HIV-1. These bacteria include Fusobacterium nucleatum (commonly present in oral cavity, and gut), Clostridium cochlearium, Eubacterium multiforme (gut), and Anaerococcus tetradius (vagina). We also clarified that butyric acid in these culture supernatants could induce histone acetylation and HIV-1 replication by inhibiting HDAC. Our observations indicate that butyric acid-producing bacteria could be involved in AIDS progression by reactivating the latent HIV provirus and, subsequently, by eliminating such bacterial infection may contribute to the prevention of the AIDS development and transmission.

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“…To test the effect of AKIP1 on natural promoter, we transfected 293 cells with the HIV-1-LTR-luc reporter gene, containing 2 B sites (34). Whereas TNF␣ stimulation induced NF-B activation by 8.6-fold, co-transfection of AKIP1 further enhanced the transcriptional activity by 1.9-fold (Fig.…”

Section: Pma (Compare Lanesmentioning

confidence: 99%

AKIP1 Enhances NF-κB-dependent Gene Expression by Promoting the Nuclear Retention and Phosphorylation of p65

Gao

Asamitsu

Hibi

et al. 2008

Journal of Biological Chemistry

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In this study, we have identified protein kinase A-interacting protein 1 (AKIP1) as a binding partner of NF-B p65 subunit, and AKIP1 enhances the NF-B-mediated gene expression. AKIP1 is a nuclear protein and known to interact with the catalytic subunit of PKA (PKAc). We identified AKIP1 by a yeast two-hybrid screen using the N terminus region of p65 as bait. The interaction between AKIP1 and p65 was confirmed by glutathione S-transferase pull-down assay in vitro and immunoprecipitation-Western blotting assay in vivo. We found that the PKAc was present in the AKIP1⅐p65 complex and enhanced the transcriptional activity of NF-B by phosphorylating p65. In a transient luciferase assay, AKIP1 cotransfection efficiently increased the transcriptional activity of NF-B induced by phorbol 12-myristate 13-acetate (PMA). When AKIP1 was knocked down by RNA interference, the PMA-mediated NF-B-dependent gene expression was abolished, indicating a physiological role of AKIP1. We found that PKAc, which is maintained in an inactive form by binding to IB␣ and NF-B in resting cells, was activated by PMA-induced signaling and could phosphorylate p65. Overexpression of AKIP1 increased the PKAc binding to p65 and enhanced the PKAc-mediated phosphorylation of p65 at Ser-276. Interestingly, this p65 phosphorylation promoted nuclear translocation of p65 and enhanced NF-B transcription. In fact, we observed that AKIP1 colocalized with p65 within the cells and appeared to retain p65 in nucleus. These findings indicate a positive role of AKIP1 in NF-B signaling and suggest a novel mechanism by which AKIP1 augments the transcriptional competence of NF-B.NF-B is an inducible transcription factor for the expression of wide variety of genes involved in immunoinflammatory responses, cell proliferation, and survival, thus playing crucial roles in the pathogenesis of many diseases including cancer, leukemia, and autoimmune diseases (1-4). NF-B exists as either a heterodimer or a homodimer, among which the p65/ p50 is the most ubiquitous heterodimer. In resting cells, NF-B dimers are sequestered in the cytoplasm through association with inhibitory proteins IBs (5). Upon treatment with NF-B inducers such as phorbol 12-myristate 13-acetate (PMA) 2 or pro-inflammatory cytokines, IB is phosphorylated and degraded through the ubiquitin/proteasome pathway, which eventually leads to the nuclear translocation of NF-B and binding to the B site of target genes (6, 7).It has been established that the phosphorylation of p65 is important for the transcriptional activity of NF-B (8 -12). The phosphorylation of p65 by the PKA catalytic subunit dramatically enhances NF-B transcriptional activity by recruiting histone acetyltransferase CBP/p300 (13-15). PKA, existing predominantly in the cytoplasm as an inactive tetramer holoenzyme in resting cells, is composed of two catalytic subunits and a homodimer of two regulatory subunits that can dissociate upon activation by cAMP (16 -20). In resting cells, PKAc is involved in the IB⅐NF-B complex present in the cytoplasm, a...

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Inhibition of Human Immunodeficiency Virus Type 1 Replication in Latently Infected Cells by a Novel IκB Kinase Inhibitor

Cited by 34 publications

References 76 publications

Involvement of Histone H3 Lysine 9 (H3K9) Methyltransferase G9a in the Maintenance of HIV-1 Latency and Its Reactivation by BIX01294

Involvement of Histone H3 Lysine 9 (H3K9) Methyltransferase G9a in the Maintenance of HIV-1 Latency and Its Reactivation by BIX01294

Reactivation of latent HIV-1 by a wide variety of butyric acid-producing bacteria

AKIP1 Enhances NF-κB-dependent Gene Expression by Promoting the Nuclear Retention and Phosphorylation of p65

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