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CNS Neuroscience & Therapeutics Jan 2023Mitochondrial-associated ER membranes (MAMs) control many cellular functions, including calcium and lipid exchange, intracellular trafficking, and mitochondrial...
INTRODUCTION
Mitochondrial-associated ER membranes (MAMs) control many cellular functions, including calcium and lipid exchange, intracellular trafficking, and mitochondrial biogenesis. The disruption of these functions contributes to neurocognitive disorders, such as spatial memory impairment and premature brain aging. Using neuronal cells, we demonstrated that HIV-1 Tat protein deregulates the mitochondria.
METHODS& RESULTS
To determine the mechanisms, we used a neuronal cell line and showed that Tat-induced changes in expression and interactions of both MAM-associated proteins and MAM tethering proteins. The addition of HIV-1 Tat protein alters expression levels of PTPIP51 and VAPB proteins in the MAM fraction but not the whole cell. Phosphorylation of PTPIP51 protein regulates its subcellular localization and function. We demonstrated that the Tat protein promotes PTPIP51 phosphorylation on tyrosine residues and prevents its binding to VAPB. Treatment of the cells with a kinase inhibitor restores the PTPIP51-VAPB interaction and overcomes the effect of Tat.
CONCLUSION
These results suggest that Tat disrupts the MAM, through the induction of PTPIP51 phosphorylation, leading to ROS accumulation, mitochondrial stress, and altered movement. Hence, we concluded that interfering in the MAM-associated cellular pathways contributes to spatial memory impairment and premature brain aging often observed in HIV-1-infected patients.
Topics: Humans; Brain; Gene Products, tat; HIV-1; Mitochondria; Protein Tyrosine Phosphatases; Endoplasmic Reticulum
PubMed: 36419337
DOI: 10.1111/cns.14011 -
Journal of Neurochemistry Jan 2003Impaired function of the brain vasculature might contribute to the development of HIV-associated dementia. For example, injury or dysfunction of brain microvascular...
Impaired function of the brain vasculature might contribute to the development of HIV-associated dementia. For example, injury or dysfunction of brain microvascular endothelial cells (BMEC) can lead to the breakdown of the blood-brain barrier (BBB) and thus allow accelerated entry of the HIV-1 virus into the CNS. Mechanisms of injury to BMEC during HIV-1 infection are not fully understood, but the viral gene product Tat may be, at least in part, responsible for this effect. Tat can be released from infected perivascular macrophages in the CNS of patients with AIDS, and thus BMEC can be directly exposed to high concentrations of this protein. To study oxidative and inflammatory mechanisms associated with Tat-induced toxicity, BMEC were exposed to increasing doses of Tat1-72, and markers of oxidative stress, as well as redox-responsive transcription factors such as nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1), were measured. Tat1-72 treatment markedly increased cellular oxidative stress, decreased levels of intracellular glutathione and activated DNA binding activity and transactivation of NF-kappaB and AP-1. To determine if Tat1-72 can stimulate inflammatory responses in brain endothelium in vivo, expression of monocyte chemoattractant protein-1 (MCP-1), an NF-kappaB and AP-1-dependent chemokine, was studied in brain tissue in mice injected with Tat1-72 into the right hippocampus. Tat1-72 markedly elevated the MCP-1 mRNA levels in brain tissue. In addition, a double immunohistochemistry study revealed that MCP-1 protein was markedly overexpressed on brain vascular endothelium. These data indicate that Tat1-72 can induce redox-related inflammatory responses both in in vitro and in vivo environments. These changes can directly lead to disruption of the BBB. Thus, Tat can play an important role in the development of detrimental vascular changes in the brains of HIV-infected patients.
Topics: Animals; Cerebrovascular Circulation; Chemokine CCL2; Clone Cells; Endothelium, Vascular; Gene Products, tat; Microcirculation; NF-kappa B; Oxidative Stress; RNA, Messenger; Swine; Transcription Factor AP-1; Transcriptional Activation; Vasculitis; tat Gene Products, Human Immunodeficiency Virus
PubMed: 12485413
DOI: 10.1046/j.1471-4159.2003.01543.x -
Microbes and Infection May 2008Monoxenic trypanosomatids, which usually are non-pathogenic in humans, have been detected in AIDS patients, but the mechanisms underlying the establishment of these...
Monoxenic trypanosomatids, which usually are non-pathogenic in humans, have been detected in AIDS patients, but the mechanisms underlying the establishment of these protozoa in HIV-1-infected individuals are poorly understood. Here we addressed the role of HIV-1 and the HIV-1 Tat protein in the replication of the monoxenic trypanosomatid Blastocrithidia culicis in HIV-1-infected primary human macrophages. We found that HIV-1 and B. culicis replication augmented almost three times in co-infected macrophages, and that Tat antiserum significantly reduced the exacerbated protozoan growth. Exposure of B. culicis only infected macrophages to Tat protein also resulted in enhanced protozoan proliferation, reaching a twofold increase at 100 ng/mL. Electron microscopy analysis revealed that B. culicis and HIV-1 co-habit the same cells, and showed protozoan dividing forms inside macrophages. Protozoan replication diminished when B. culicis only infected macrophages were treated with Tat protein in the presence of anti-TGF-beta1 antibodies, suggesting a participation of this cytokine in the augmentation of protozoan multiplication. In fact, exogenous TGF-beta1 promoted the trypanosomatid replication in macrophages. Overall, our results suggest that HIV-1 infection deactivates the macrophage microbicidal activity, permitting the survival and multiplication of an otherwise non-pathogenic protozoan in these cells, a process partially mediated by Tat protein, via TGF-beta1 secretion.
Topics: Animals; HIV Infections; HIV-1; Humans; Macrophages; Transforming Growth Factor beta1; Trypanosomatina; tat Gene Products, Human Immunodeficiency Virus
PubMed: 18457978
DOI: 10.1016/j.micinf.2008.02.014 -
Clinical Microbiology Reviews Apr 2002The Tat protein of human immunodeficiency virus (HIV) type 1 is a transactivator of viral gene expression that is required for virus replication and spread. Moreover,... (Review)
Review
The Tat protein of human immunodeficiency virus (HIV) type 1 is a transactivator of viral gene expression that is required for virus replication and spread. Moreover, Tat is released by acutely HIV-infected cells via a leaderless secretory pathway and in a biologically active form that exerts effects on both HIV-infected and uninfected cells from different organs and systems. This review focuses on the activities of extracellular Tat protein on endothelial cells, on angiogenesis, and on the pathogenesis of AIDS-associated angioproliferative diseases such as Kaposi's sarcoma. In particular, we discuss results from different groups indicating that Tat mimics the proangiogenic activities of extracellular matrix molecules and that it enhances the effects of angiogenic factors.
Topics: AIDS-Related Opportunistic Infections; Animals; Gene Products, tat; HIV Infections; HIV-1; Humans; Mice; Neovascularization, Pathologic; Sarcoma, Kaposi; tat Gene Products, Human Immunodeficiency Virus
PubMed: 11932235
DOI: 10.1128/CMR.15.2.310-326.2002 -
RNA Biology Jul 2017The trans-activator Tat protein of HIV-1 belongs to the large family of intrinsically disordered proteins (IDPs), and is known to recruit various host proteins for the...
The trans-activator Tat protein of HIV-1 belongs to the large family of intrinsically disordered proteins (IDPs), and is known to recruit various host proteins for the transactivation of viral RNA synthesis. Tat protein interacts with the transactivator response RNA (TAR RNA), exhibiting RNA chaperone activities for structural rearrangement of interacting RNAs. Here, considering that Tat-TAR RNA interaction is mutually cooperative, we examined the potential role of TAR RNA as Chaperna - RNA that provides chaperone function to proteins - for the folding of HIV-1 Tat. Using EGFP fusion as an indirect indicator for folding status, we monitored Tat-EGFP folding in HeLa cells via time-lapse fluorescence microscopy. The live cell imaging showed that the rate and the extent of folding of Tat-EGFP were stimulated by TAR RNA. The purified Tat-EGFP was denatured and the fluorescence was monitored in vitro under renaturation condition. The fluorescence was significantly increased by TAR RNA, and the mutations in TAR RNA that affected the interaction with Tat protein failed to promote Tat refolding. The results suggest that TAR RNA stabilizes Tat as unfolded, but prevents it from misfolding, and maintaining its folding competence for interaction with multiple host factors toward its transactivation. The Chaperna function of virally encoded RNA in establishing proteome link at the viral-host interface provides new insights to as yet largely unexplored RNA mediated protein folding in normal and dysregulated cellular metabolism.
Topics: Base Sequence; Green Fluorescent Proteins; HIV Long Terminal Repeat; HIV-1; HeLa Cells; Humans; Protein Binding; Protein Folding; Recombinant Fusion Proteins; Solubility; Subcellular Fractions; Time Factors; tat Gene Products, Human Immunodeficiency Virus
PubMed: 28418268
DOI: 10.1080/15476286.2017.1311455 -
Retrovirology Jul 2005HIV-1 Tat protein recruits human positive transcription elongation factor P-TEFb, consisting of CDK9 and cyclin T1, to HIV-1 transactivation response (TAR) RNA. CDK9 is...
BACKGROUND
HIV-1 Tat protein recruits human positive transcription elongation factor P-TEFb, consisting of CDK9 and cyclin T1, to HIV-1 transactivation response (TAR) RNA. CDK9 is maintained in dephosphorylated state by TFIIH and undergo phosphorylation upon the dissociation of TFIIH. Thus, dephosphorylation of CDK9 prior to its association with HIV-1 preinitiation complex might be important for HIV-1 transcription. Others and we previously showed that protein phosphatase-2A and protein phosphatase-1 regulates HIV-1 transcription. In the present study we analyze relative contribution of PP2A and PP1 to dephosphorylation of CDK9 and to HIV-1 transcription in vitro and in vivo.
RESULTS
In vitro, PP2A but not PP1 dephosphorylated autophosphorylated CDK9 and reduced complex formation between P-TEFb, Tat and TAR RNA. Inhibition of PP2A by okadaic acid inhibited basal as well as Tat-induced HIV-1 transcription whereas inhibition of PP1 by recombinant nuclear inhibitor of PP1 (NIPP1) inhibited only Tat-induced transcription in vitro. In cultured cells, low concentration of okadaic acid, inhibitory for PP2A, only mildly inhibited Tat-induced HIV-1 transcription. In contrast Tat-mediated HIV-1 transcription was strongly inhibited by expression of NIPP1. Okadaic acid induced phosphorylation of endogenous as well transiently expressed CDK9, but this induction was not seen in the cells expressing NIPP1. Also the okadaic acid did not induce phosphorylation of CDK9 with mutation of Thr 186 or with mutations in Ser-329, Thr-330, Thr-333, Ser-334, Ser-347, Thr-350, Ser-353, and Thr-354 residues involved in autophosphorylation of CDK9.
CONCLUSION
Our results indicate that although PP2A dephosphorylates autophosphorylated CDK9 in vitro, in cultured cells PP1 is likely to dephosphorylate CDK9 and contribute to the regulation of activated HIV-1 transcription.
Topics: Animals; COS Cells; Chlorocebus aethiops; Cyclin-Dependent Kinase 9; Gene Products, tat; HIV Long Terminal Repeat; HIV-1; Humans; Phosphoprotein Phosphatases; Phosphorylation; Protein Phosphatase 1; Protein Phosphatase 2; Transcription, Genetic; tat Gene Products, Human Immunodeficiency Virus
PubMed: 16048649
DOI: 10.1186/1742-4690-2-47 -
Proceedings of the National Academy of... Nov 1998In cells infected with HIV type 1 (HIV-1), the integrated viral promoter is present in a chromatin-bound conformation and is transcriptionally silent in the absence of...
In cells infected with HIV type 1 (HIV-1), the integrated viral promoter is present in a chromatin-bound conformation and is transcriptionally silent in the absence of stimulation. The HIV-1 Tat protein binds to a stem-loop structure at the 5' end of viral mRNA and relieves this inhibition by inducing a remodeling of the nucleosome arrangement downstream of the transcription-initiation site. Here we show that Tat performs this activity by recruiting to the viral long terminal repeat (LTR) the transcriptional coactivator p300 and the closely related CREB-binding protein (CBP), having histone acetyltransferase (HAT) activity. Tat associates with HAT activity in human nuclear extracts and binds to p300 and CBP both in vitro and in vivo. Integrity of the basic domain of Tat is essential for this interaction. By a quantitative chromatin immunoprecipitation assay we show that the delivery of recombinant Tat induces the association of p300 and CBP with the chromosomally integrated LTR promoter. Expression of human p300 in both human and rodent cells increases the levels of Tat transactivation of the integrated LTR. These results reinforce the evidence that p300 and CBP have a pivotal function at both cellular and viral promoters and demonstrate that they also can be recruited by an RNA-targeted activator. Additionally, these findings have important implications for the understanding of the mechanisms of HIV-1 latency and reactivation.
Topics: CREB-Binding Protein; Cell Line; Gene Products, tat; HIV Infections; HIV Long Terminal Repeat; HIV-1; Histone Acetyltransferases; Humans; Nuclear Proteins; Nuclear Receptor Coactivator 3; Trans-Activators; Virus Integration; tat Gene Products, Human Immunodeficiency Virus
PubMed: 9811832
DOI: 10.1073/pnas.95.23.13519 -
Journal of Microbiology (Seoul, Korea) Dec 2004The human immunodeficiency virus type 1 (HIV-1) Tat protein transduction domain (PTD), which contains rich arginine and lysine residues, is responsible for the highly...
The human immunodeficiency virus type 1 (HIV-1) Tat protein transduction domain (PTD), which contains rich arginine and lysine residues, is responsible for the highly efficient transduction of protein through the plasma membrane. In addition, it can be secreted from infected cells and has the ability to enter neighboring cells. When the PTD of Tat is fused to proteins and exogenously added to cells, the fusion protein can cross plasma membranes. Recent reports indicate that the endogenously expressed Tat fusion protein can demonstrate biodistribution of several proteins. However, intercellular transport and protein transduction have not been observed in some studies. Therefore, this study examined the intercellular transport and protein transduction of the Tat protein. The results showed no evidence of intercellular transport (biodistribution) in a cell culture. Instead, the Tat fusion peptides were found to have a significant effect on the transduction and intercellular localization properties. This suggests that the HIV-1 PTD passes through the plasma membrane in one direction.
Topics: Animals; Baculoviridae; Cell Line; Cell Line, Tumor; Cell Membrane; Gene Products, tat; Genetic Vectors; Green Fluorescent Proteins; HIV-1; Intracellular Space; Protein Structure, Tertiary; Protein Transport; Recombinant Proteins; Transduction, Genetic; tat Gene Products, Human Immunodeficiency Virus
PubMed: 15650690
DOI: No ID Found -
The EMBO Journal Jun 2002The human immunodeficiency virus (HIV) Tat protein plays an essential role in promoting efficient transcriptional elongation of viral transcripts. We report that the...
The human immunodeficiency virus (HIV) Tat protein plays an essential role in promoting efficient transcriptional elongation of viral transcripts. We report that the transcriptional co-activator PCAF and Tat interact and synergize to activate the HIV promoter. The binding of Tat and PCAF in vitro and in vivo is dependent on the acetylated state of Lys50 of Tat and on the PCAF bromodomain. Structural analysis of the acetylated Tat peptide bound to the PCAF bromodomain defined amino acids Y47 and R53 in Tat and V763, Y802, and Y809 in PCAF as critical interaction points between the two proteins. Mutation of each of these residues in either Tat or PCAF inhibited in a cumulative manner the Tat-PCAF interaction in vitro and in vivo, and abrogated the synergistic activation of the HIV promoter by both proteins. These observations demonstrate that acetylation of Tat establishes a novel protein-protein interaction domain at the surface of Tat that is necessary for the transcriptional activation of the HIV promoter.
Topics: Acetyltransferases; Blotting, Western; Cell Line; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Gene Products, tat; Green Fluorescent Proteins; HeLa Cells; Histone Acetyltransferases; Humans; Luciferases; Luminescent Proteins; Lysine; Peptides; Plasmids; Precipitin Tests; Promoter Regions, Genetic; Protein Binding; Protein Structure, Tertiary; Recombinant Proteins; Saccharomyces cerevisiae Proteins; Transcription, Genetic; Transcriptional Activation; Transfection
PubMed: 12032084
DOI: 10.1093/emboj/21.11.2715 -
Journal of Virology Nov 1998Chemokine receptors CCR5 and CXCR4 are the primary fusion coreceptors utilized for CD4-mediated entry by macrophage (M)- and T-cell line (T)-tropic human...
Chemokine receptors CCR5 and CXCR4 are the primary fusion coreceptors utilized for CD4-mediated entry by macrophage (M)- and T-cell line (T)-tropic human immunodeficiency virus type 1 (HIV-1) strains, respectively. Here we demonstrate that HIV-1 Tat protein, a potent viral transactivator shown to be released as a soluble protein by infected cells, differentially induced CXCR4 and CCR5 expression in peripheral blood mononuclear cells. CCR3, a less frequently used coreceptor for certain M-tropic strains, was also induced. CXCR4 was induced on both lymphocytes and monocytes/macrophages, whereas CCR5 and CCR3 were induced on monocytes/macrophages but not on lymphocytes. The pattern of chemokine receptor induction by Tat was distinct from that by phytohemagglutinin. Moreover, Tat-induced CXCR4 and CCR5 expression was dose dependent. Monocytes/macrophages were more susceptible to Tat-mediated induction of CXCR4 and CCR5 than lymphocytes, and CCR5 was more readily induced than CXCR4. The concentrations of Tat effective in inducing CXCR4 and CCR5 expression were within the picomolar range and close to the range of extracellular Tat observed in sera from HIV-1-infected individuals. The induction of CCR5 and CXCR4 expression correlated with Tat-enhanced infectivity of M- and T-tropic viruses, respectively. Taken together, our results define a novel role for Tat in HIV-1 pathogenesis that promotes the infectivity of both M- and T-tropic HIV-1 strains in primary human leukocytes, notably in monocytes/macrophages.
Topics: Base Sequence; Cells, Cultured; DNA Primers; Dose-Response Relationship, Drug; Gene Expression; Gene Products, tat; HIV Infections; HIV-1; Humans; Macrophages; Monocytes; Phytohemagglutinins; Receptors, CCR3; Receptors, CCR5; Receptors, CXCR4; Receptors, Chemokine; Recombinant Proteins; T-Lymphocytes; tat Gene Products, Human Immunodeficiency Virus
PubMed: 9765440
DOI: 10.1128/JVI.72.11.8952-8960.1998