scholarly journals Modulation of the Brd4/P-TEFb Interaction by the Human T-Lymphotropic Virus Type 1 Tax Protein

2007 ◽  
Vol 81 (20) ◽  
pp. 11179-11186 ◽  
Author(s):  
Won-Kyung Cho ◽  
Meisheng Zhou ◽  
Moon Kyoo Jang ◽  
Keven Huang ◽  
Soo-Jin Jeong ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Virus Type ◽  
Active Form ◽  
Viral Gene ◽  
Dependent Manner ◽  
Binding Studies ◽  
T Lymphotropic Virus ◽  
Cyclin T1

ABSTRACT Positive transcription elongation factor (P-TEFb), which is composed of CDK9 and cyclin T1, plays an important role in cellular and viral gene expression. Our lab has recently demonstrated that P-TEFb is required for Tax transactivation of the viral long terminal repeat (LTR). P-TEFb is found in two major complexes: the inactive form, which is associated with inhibitory subunits 7SK snRNA and HEXIM1, and the active form, which is associated with, at least in part, Brd4. In this study, we analyzed the effect of Brd4 on human T-lymphotropic virus type 1 (HTLV-1) transcription. Overexpression of Brd4 repressed Tax transactivation of the HTLV-1 LTR in a dose-dependent manner. In vitro binding studies suggest that Tax and Brd4 compete for binding to P-TEFb through direct interaction with cyclin T1. Tax interacts with cyclin T1 amino acids 426 to 533, which overlaps the region responsible for Brd4 binding. In vivo, overexpression of Tax decreased the amount of 7SK snRNA associated with P-TEFb and stimulates serine 2 phosphorylation of the RNA polymerase II carboxyl-terminal domain, suggesting that Tax regulates the functionality of P-TEFb. Our results suggest the possibility that Tax may compete and functionally substitute for Brd4 in P-TEFb regulation.

scholarly journals Human T-Lymphotropic Virus Type 1 Tax Protein Complexes with P-TEFb and Competes for Brd4 and 7SK snRNP/HEXIM1 Binding

2010 ◽  
Vol 84 (24) ◽  
pp. 12801-12809 ◽  
Author(s):  
Won-Kyung Cho ◽  
Moon Kyoo Jang ◽  
Keven Huang ◽  
Cynthia A. Pise-Masison ◽  
John N. Brady
Keyword(s):  
Molecular Weight ◽  
Rna Polymerase Ii ◽  
Virus Type ◽  
T Lymphotropic Virus ◽  
Cyclin T1 ◽  
Tax Protein ◽  
In Cells

ABSTRACT Positive transcription elongation factor b (P-TEFb) plays an important role in stimulating RNA polymerase II elongation for viral and cellular gene expression. P-TEFb is found in cells in either an active, low-molecular-weight (LMW) form or an inactive, high-molecular-weight (HMW) form. We report here that human T-lymphotropic virus type 1 (HTLV-1) Tax interacts with the cyclin T1 subunit of P-TEFb, forming a distinct Tax/P-TEFb LMW complex. We demonstrate that Tax can play a role in regulating the amount of HMW complex present in the cell by decreasing the binding of 7SK snRNP/HEXIM1 to P-TEFb. This is seen both in vitro using purified Tax protein and in vivo in cells transduced with Tax expression constructs. Further, we find that a peptide of cyclin T1 spanning the Tax binding domain inhibits the ability of Tax to disrupt HMW P-TEFb complexes. These results suggest that the direct interaction of Tax with cyclin T1 can dissociate P-TEFb from the P-TEFb/7SK snRNP/HEXIM1 complex for activation of the viral long terminal repeat (LTR). We also show that Tax competes with Brd4 for P-TEFb binding. Chromatin immunoprecipitation (ChIP) assays demonstrated that Brd4 and P-TEFb are associated with the basal HTLV-1 LTR, while Tax and P-TEFb are associated with the activated template. Furthermore, the knockdown of Brd4 by small interfering RNA (siRNA) activates the HTLV-1 LTR promoter, which results in an increase in viral expression and production. Our studies have identified Tax as a regulator of P-TEFb that is capable of affecting the balance between its association with the large inactive complex and the small active complex.

scholarly journals Tax Interacts with P-TEFb in a Novel Manner To Stimulate Human T-Lymphotropic Virus Type 1 Transcription

2006 ◽  
Vol 80 (10) ◽  
pp. 4781-4791 ◽  
Author(s):  
Meisheng Zhou ◽  
Hanxin Lu ◽  
Hyeon Park ◽  
Jaime Wilson-Chiru ◽  
Rebecca Linton ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Virus Type ◽  
Protein Interactions ◽  
Elongation Factor ◽  
Protein Protein Interactions ◽  
T Lymphotropic Virus ◽  
Histone Deacetylase 1

ABSTRACT Human T-lymphotropic virus type 1 (HTLV-1) encodes a transcriptional activator, Tax, whose function is essential for viral transcription and replication. Tax transactivates the viral long-terminal repeat through a series of protein-protein interactions which facilitate CREB and CBP/p300 binding. In addition, Tax dissociates transcription repressor histone deacetylase 1 interaction with the CREB response element. The subsequent events through which Tax interacts and communicates with RNA polymerase II and cyclin-dependent kinases (CDKs) are not clearly understood. Here we present evidence that Tax recruits positive transcription elongation factor b (P-TEFb) (CDK9/cyclin T1) to the viral promoter. This recruitment likely involves protein-protein interactions since Tax associates with P-TEFb in vitro as demonstrated by glutathione S-transferase fusion protein pull-down assays and in vivo as shown by coimmunoprecipitation assays. Functionally, small interfering RNA directed toward CDK9 inhibited Tax transactivation in transient assays. Consistent with these findings, the depletion of CDK9 from nuclear extracts inhibited Tax transactivation in vitro. Reconstitution of the reaction with wild-type P-TEFb, but not a kinase-dead mutant, recovered HTLV-1 transcription. Moreover, the addition of the CDK9 inhibitor flavopiridol blocked Tax transactivation in vitro and in vivo. Interestingly, we found that Tax regulates CDK9 kinase activity through a novel autophosphorylation pathway.

The Role of Human T-Lymphotropic Virus Type 1 (HTLV-1)-Infected Dendritic Cells in the Development of HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis

1999 ◽  
Vol 73 (6) ◽  
pp. 4575-4581 ◽  
Author(s):  
Masahiko Makino ◽  
Satoshi Shimokubo ◽  
Shin-Ichi Wakamatsu ◽  
Shuji Izumo ◽  
Masanori Baba
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Virus Type ◽  
Spastic Paraparesis ◽  
Tropical Spastic Paraparesis ◽  
Normal Donor ◽  
Dependent Manner ◽  
T Lymphotropic Virus

ABSTRACT The development of human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is closely associated with the activation of T cells which are HTLV-1 specific but may cross-react with neural antigens (Ags). Immature dendritic cells (DCs), differentiated from normal donor monocytes by using recombinant granulocyte-macrophage colony-stimulating factor and recombinant interleukin-4, were pulsed with HTLV-1 in vitro. The pulsed DCs contained HTLV-1 proviral DNA and expressed HTLV-1 Gag Ag on their surface 6 days after infection. The DCs matured by lipopolysaccharides stimulated autologous CD4+ T cells and CD8+ T cells in a viral dose-dependent manner. However, the proliferation level of CD4+ T cells was five- to sixfold higher than that of CD8+ T cells. In contrast to virus-infected DCs, DCs pulsed with heat-inactivated virions activated only CD4+ T cells. To clarify the role of DCs in HAM/TSP development, monocytes from patients were cultured for 4 days in the presence of the cytokines. The expression of CD86 Ag on DCs was higher and that of CD1a Ag was more down-regulated than in DCs generated from normal monocytes. DCs from two of five patients expressed HTLV-1 Gag Ag. Furthermore, both CD4+ and CD8+ T cells from the patients were greatly stimulated by contact with autologous DCs pulsed with inactivated viral Ag as well as HTLV-1-infected DCs. These results suggest that DCs are susceptible to HTLV-1 infection and that their cognate interaction with T cells may contribute to the development of HAM/TSP.

scholarly journals Wogonin inhibits in vitro herpes simplex virus type 1 and 2 infection through modulating cellular NF-κB and MAPK pathways

2020 ◽  
Author(s):  
Ying Chu ◽  
Xiaowen Lv ◽  
Longfeng Zhang ◽  
Xingli Fu ◽  
Siwei Song ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Dependent Manner ◽  
Mapk Pathways ◽  
Simplex Virus ◽  
Hsv 1

Abstract Background: Wogonin, a naturally flavonoid-like chemical compounds, has exhibited anti-inflammatory, anti-tumor, anti-viral, neuroprotective, and anxiolytic effects through modulating a variety of signaling pathways including PI3K-Akt, p53, nuclear factor κB (NF-κB) and mitogen-activated protein kinases (MAPK). In this study, its antiviral effect against herpes simplex virus type 1 and 2 (HSV-1 and HSV-2) replication has been investigated. Results: The evidence showed that wogonin could suppress HSV-1 and -2-induced cytopathogenic effect (CPE) and reduce viral RNA transcription, protein synthesis, and titers of infectious virion particles formation in a dose-dependent manner. Time-of-drug-addition assay demonstrated that wogonin acted as a viral post-entry inhibitor . It was also found that wogonin significantly reduced HSV-induced NF-κB and MAPK pathways activation, which were proved to be significant for viral replication previously. Conclusions: Our results suggested that anti-herpes effect of wogonin may be mediated by its modulation of cellular NF-κB and JNK/p38 MAPK pathways, implying its potential application as an anti-HSV agent.

scholarly journals Human immunodeficiency virus type 1 Tat prevents dephosphorylation of Sp1 by TCF-4 in astrocytes

2006 ◽  
Vol 87 (6) ◽  
pp. 1613-1623 ◽  
Author(s):  
Andrea Rossi ◽  
Ruma Mukerjee ◽  
Pasquale Ferrante ◽  
Kamel Khalili ◽  
Shohreh Amini ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Viral Gene ◽  
Physical Interaction ◽  
Rich Domain ◽  
Immunodeficiency Virus ◽  
Hiv 1

Previous examination of the effect of TCF-4 on transcription of the human immunodeficiency virus type 1 (HIV-1) promoter in human astrocytic cells found that TCF-4 affects the HIV-1 promoter through the GC-rich domain (nt −80 to nt −68). Here, the physical interaction and a functional consequence of TCF4–Sp1 contact were characterized. It was shown that expression of TCF-4 in U-87 MG (human astrocytic) cells decreased basal and Sp1-mediated transcription of the HIV-1 promoter. Results from a GST pull-down assay, as well as combined immunoprecipitation and Western blot analysis of protein extracts from U-87 MG cells, revealed an interaction of Sp1 with TCF-4. Using in vitro protein chromatography, the region of Sp1 that contacts TCF-4 was mapped to aa 266–350. It was also found that, in cell-free extracts, TCF-4 prevented dsDNA-dependent protein kinase (DNA-PK)-mediated Sp1 phosphorylation. Surprisingly, TCF-4 failed to decrease Sp1-mediated transcription of the HIV-1 long terminal repeat (LTR) and Sp1 phosphorylation in cells expressing HIV-1 Tat. Results from immunoprecipitation/Western blotting demonstrated that TCF-4 lost its ability to interact with Sp1, but not with Tat, in Tat-transfected cells. Taken together, these findings suggest that activity at the HIV-1 promoter is influenced by phosphorylation of Sp1, which is affected by Tat and DNA-PK. Interactions among TCF-4, Sp1 and/or Tat may determine the level of viral gene transcription in human astrocytic cells.

scholarly journals XBP-1, a Novel Human T-Lymphotropic Virus Type 1 (HTLV-1) Tax Binding Protein, Activates HTLV-1 Basal and Tax-Activated Transcription

2008 ◽  
Vol 82 (9) ◽  
pp. 4343-4353 ◽  
Author(s):  
Sebastian C. Y. Ku ◽  
Jialing Lee ◽  
Joanne Lau ◽  
Meera Gurumurthy ◽  
Raymond Ng ◽  
...  
Keyword(s):  
Virus Type ◽  
Binding Protein ◽  
Host Cells ◽  
Mrna Levels ◽  
Transcription Activator ◽  
Basic Leucine Zipper ◽  
T Lymphotropic Virus ◽  
In Cells

ABSTRACT X-box binding protein 1 (XBP-1), a basic leucine zipper transcription factor, plays a key role in the cellular unfolded protein response (UPR). There are two XBP-1 isoforms in cells, spliced XBP-1S and unspliced XBP-1U. XBP-1U has been shown to bind to the 21-bp Tax-responsive element of the human T-lymphotropic virus type 1 (HTLV-1) long terminal repeat (LTR) in vitro and transactivate HTLV-1 transcription. Here we identify XBP-1S as a transcription activator of HTLV-1. Compared to XBP-1U, XBP-1S demonstrates stronger activating effects on both basal and Tax-activated HTLV-1 transcription in cells. Our results show that both XBP-1S and XBP-1U interact with Tax and bind to the HTLV-1 LTR in vivo. In addition, elevated mRNA levels of the gene for XBP-1 and several UPR genes were detected in the HTLV-1-infected C10/MJ and MT2 T-cell lines, suggesting that HTLV-1 infection may trigger the UPR in host cells. We also identify Tax as a positive regulator of the expression of the gene for XBP-1. Activation of the UPR by tunicamycin showed no effect on the HTLV-1 LTR, suggesting that HTLV-1 transcription is specifically regulated by XBP-1. Collectively, our study demonstrates a novel host-virus interaction between a cellular factor XBP-1 and transcriptional regulation of HTLV-1.

scholarly journals Transient Induction of Cyclin T1 during Human Macrophage Differentiation Regulates Human Immunodeficiency Virus Type 1 Tat Transactivation Function

2002 ◽  
Vol 76 (21) ◽  
pp. 10579-10587 ◽  
Author(s):  
Li-Ying Liou ◽  
Christine H. Herrmann ◽  
Andrew P. Rice
Keyword(s):  
Human Immunodeficiency Virus ◽  
Protein Expression ◽  
Rna Polymerase Ii ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Human Macrophage ◽  
Macrophage Differentiation ◽  
Cyclin T1 ◽  
Immunodeficiency Virus

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) Tat protein is essential for viral replication and stimulates transcription of the integrated provirus by recruiting the kinase complex TAK/P-TEFb, composed of cyclin T1 (CycT1) and Cdk9, to the viral TAR RNA element. TAK/P-TEFb phosphorylates the RNA polymerase II complex and stimulates transcriptional elongation. In this report, we investigated the regulation of TAK/P-TEFb in primary human macrophages, a major target cell of HIV infection. While Cdk9 levels remained constant, CycT1 protein expression in freshly isolated monocytes was very low, increased early during macrophage differentiation, and, unexpectedly, decreased to very low levels after about 1 week in culture. The kinase activity of TAK/P-TEFb paralleled the changes in CycT1 protein expression. RNA analysis indicated that the transient induction of CycT1 protein expression involves a posttranscriptional mechanism. In transient transfection assays, the ability of Tat to transactivate the HIV long terminal repeat (LTR) in the late differentiated macrophages was greatly diminished relative to its ability to transactivate the HIV LTR in early differentiated cells, strongly suggesting that CycT1 is limiting for Tat function in late differentiated macrophages. Interestingly, lipopolysaccharide, a component of the cell wall of gram-negative bacteria, reinduced CycT1 expression late in macrophage differentiation. These results raise the possibility that regulation of CycT1 expression may be involved in establishing latent infection in macrophages and that opportunistic infection may reactivate the virus by inducing CycT1 expression.

scholarly journals Functional Role of pX Open Reading Frame II of Human T-Lymphotropic Virus Type 1 in Maintenance of Viral Loads In Vivo

2000 ◽  
Vol 74 (3) ◽  
pp. 1094-1100 ◽  
Author(s):  
Joshua T. Bartoe ◽  
Björn Albrecht ◽  
Nathaniel D. Collins ◽  
Michael D. Robek ◽  
Lee Ratner ◽  
...  
Keyword(s):  
Virus Type ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Peripheral Blood Mononuclear ◽  
T Lymphotropic Virus ◽  
Viral Loads

ABSTRACT Human T-lymphotropic virus type 1 (HTLV-1) causes adult T-cell leukemia/lymphoma and is associated with a variety of immune-mediated disorders. The role of four open reading frames (ORFs), located between env and the 3′ long terminal repeat of HTLV-1, in mediating disease is not entirely clear. By differential splicing, ORF II encodes two proteins, p13II and p30II, both of which have not been functionally defined. p13II localizes to mitochondria and may alter the configuration of the tubular network of this cellular organelle. p30II localizes to the nucleolus and shares homology with the transcription factors Oct-1 and -2, Pit-1, and POU-M1. Both p13II and p30II are dispensable for infection and immortalization of primary human and rabbit lymphocytes in vitro. To test the role of ORF II gene products in vivo, we inoculated rabbits with lethally irradiated cell lines expressing the wild-type molecular clone of HTLV-1 (ACH.1) or a clone containing selected mutations in ORF II (ACH.30/13.1). ACH.1-inoculated animals maintained higher HTLV-1-specific antibody titers than animals inoculated with ACH.30/13.1. Viral p19 antigen was transiently detected in ex vivo cultures of peripheral blood mononuclear cells (PBMC) from only two ACH.30/13.1-inoculated rabbits, while PBMC cultures from all ACH.1-inoculated rabbits routinely produced p19 antigen. In only three of six animals exposed to the ACH.p30II/p13IIclone could provirus be consistently PCR amplified from extracted PBMC DNA and quantitative competitive PCR showed the proviral loads in PBMC from ACH.p30II/p13II-infected rabbits to be dramatically lower than the proviral loads in rabbits exposed to ACH. Our data indicate selected mutations in pX ORF II diminish the ability of HTLV-1 to maintain high viral loads in vivo and suggest an important function for p13II and p30II in viral pathogenesis.

scholarly journals CDK13, a New Potential Human Immunodeficiency Virus Type 1 Inhibitory Factor Regulating Viral mRNA Splicing

2008 ◽  
Vol 82 (14) ◽  
pp. 7155-7166 ◽  
Author(s):  
Reem Berro ◽  
Caitlin Pedati ◽  
Kylene Kehn-Hall ◽  
Weilin Wu ◽  
Zachary Klase ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Rna Polymerase Ii ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Virus Production ◽  
Mrna Splicing ◽  
Viral Gene ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) Tat is a 14-kDa viral protein that acts as a potent transactivator by binding to the transactivation-responsive region, a structured RNA element located at the 5′ end of all HIV-1 transcripts. Tat transactivates viral gene expression by inducing the phosphorylation of the C-terminal domain of RNA polymerase II through several Tat-activated kinases and by recruiting chromatin-remodeling complexes and histone-modifying enzymes to the HIV-1 long terminal repeat. Histone acetyltransferases, including p300 and hGCN5, not only acetylate histones but also acetylate Tat at lysine positions 50 and 51 in the arginine-rich motif. Acetylated Tat at positions 50 and 51 interacts with a specialized protein module, the bromodomain, and recruits novel factors having this particular domain, such as P/CAF and SWI/SNF. In addition to having its effect on transcription, Tat has been shown to be involved in splicing. In this study, we demonstrate that Tat interacts with cyclin-dependent kinase 13 (CDK13) both in vivo and in vitro. We also found that CDK13 increases HIV-1 mRNA splicing and favors the production of the doubly spliced protein Nef. In addition, we demonstrate that CDK13 acts as a possible restriction factor, in that its overexpression decreases the production of the viral proteins Gag and Env and subsequently suppresses virus production. Using small interfering RNA against CDK13, we show that silencing of CDK13 leads to a significant increase in virus production. Finally, we demonstrate that CDK13 mediates its effect on splicing through the phosphorylation of ASF/SF2.

scholarly journals The Human Polycomb Group EED Protein Interacts with the Integrase of Human Immunodeficiency Virus Type 1

2003 ◽  
Vol 77 (23) ◽  
pp. 12507-12522 ◽  
Author(s):  
Sébastien Violot ◽  
Saw See Hong ◽  
Dina Rakotobe ◽  
Caroline Petit ◽  
Bernard Gay ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Binding Site ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Dependent Manner ◽  
Immunodeficiency Virus ◽  
Infected Cells ◽  
Hiv 1

ABSTRACT Human EED, a member of the superfamily of WD-40 repeat proteins and of the Polycomb group proteins, has been identified as a cellular partner of the human immunodeficiency virus type 1 (HIV-1) matrix (MA) protein (R. Peytavi et al., J. Biol. Chem. 274:1635-1645, 1999). In the present study, EED was found to interact with HIV-1 integrase (IN) both in vitro and in vivo in yeast. In vitro, data from mutagenesis studies, pull-down assays, and phage biopanning suggested that EED-binding site(s) are located in the C-terminal domain of IN, between residues 212 and 264. In EED, two putative discrete IN-binding sites were mapped to its N-terminal moiety, at a distance from the MA-binding site, but EED-IN interaction also required the integrity of the EED last two WD repeats. EED showed an apparent positive effect on IN-mediated DNA integration reaction in vitro, in a dose-dependent manner. In situ analysis by immunoelectron microscopy (IEM) of cellular distribution of IN and EED in HIV-1-infected cells (HeLa CD4+ cells or MT4 lymphoid cells) showed that IN and EED colocalized in the nucleus and near nuclear pores, with maximum colocalization events occurring at 6 h postinfection (p.i.). Triple colocalizations of IN, EED, and MA were also observed in the nucleoplasm of infected cells at 6 h p.i., suggesting the ocurrence of multiprotein complexes involving these three proteins at early steps of the HIV-1 virus life cycle. Such IEM patterns were not observed with a noninfectious, envelope deletion mutant of HIV-1.

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