Association of HIV-1 Tat with the cellular protein, Purα, is mediated by RNA

Gallia, Gary L.; Darbinian, Nune; Tretiakova, Anna P.; Ansari, Sameer A.; Rappaport, Jay; Brady, John; Wortman, Margaret J.; Johnson, Edward M.; Khalili, Kamel

doi:10.1073/pnas.96.20.11572

Cited by 53 publications

(60 citation statements)

References 29 publications

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“…10 and 11). Such an observation is quite interesting given the precedent that human Pur␣ can bind to other transcriptionally relevant proteins such as pRb (26,39), MyEF-2 (40), JC virus T-antigen (41), HIV-1 Tat protein (42,43), YB-1 (19), E2F-1 (44), Sp1 (45), and hnRNP K (46). Detection of Pur␣, Pur␤, and MSY1 co-immunoprecipitating with TEF-1 (and SRF) provides a strong biochemical basis for future studies aimed at evaluating whether such protein-protein associations might interfere with the ability of TEF-1 and SRF to interact with each other, with coactivating factors, or with components of the general transcription factor complex.…”

Section: Discussionmentioning

confidence: 99%

Cryptic MCAT Enhancer Regulation in Fibroblasts and Smooth Muscle Cells

Carlini

Getz

Strauch

et al. 2002

Journal of Biological Chemistry

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An asymmetric polypurine-polypyrimidine cis-element located in the 5 region of the mouse vascular smooth muscle ␣-actin gene serves as a binding site for multiple proteins with specific affinity for either singleor double-stranded DNA. Here, we test the hypothesis that single-stranded DNA-binding proteins are responsible for preventing a cryptic MCAT enhancer centered within this element from cooperating with a nearby serum response factor-interacting CArG motif to transactivate the minimal promoter in fibroblasts and smooth muscle cells. DNA binding studies revealed that the core MCAT sequence mediates binding of transcription enhancer factor-1 to the double-stranded polypurine-polypyrimidine element while flanking nucleotides account for interaction of Pur␣ and Pur␤ with the purine-rich strand and MSY1 with the complementary pyrimidine-rich strand. Mutations that selectively impaired high affinity single-stranded DNA binding by fibroblast or smooth muscle cell-derived Pur␣, Pur␤, and MSY1 in vitro, released the cryptic MCAT enhancer from repression in transfected cells. Additional experiments indicated that Pur␣, Pur␤, and MSY1 also interact specifically, albeit weakly, with double-stranded DNA and with transcription enhancer factor-1. These results are consistent with two plausible models of cryptic MCAT enhancer regulation by Pur␣, Pur␤, and MSY1 involving either competitive single-stranded DNA binding or masking of MCAT-bound transcription enhancer factor-1.Current models of transcriptional repression take into account the ability of negative-acting factors to function in multiple capacities with or without DNA binding specificity (1, 2). For example, evidence exists that certain activator proteins can simply be masked or sequestered by protein binding partners that do not interact with DNA directly, as in the case of retinoblastoma tumor suppressor protein-mediated repression of the E2F family of trans-activators (3). Alternatively, a repressor protein may bind and displace components of the basal transcription machinery as in the case of Drosophila evenskipped-mediated repression of the Adh proximal promoter (4). Recent work suggests that some repressors can participate, indirectly, in modifying chromatin structure by binding activator sites and recruiting specific histone deacetylases to silence genes through histone modification. This scenario is implicated in repression of E-box motifs through Mad/Max-mediated recruitment of histone deacetylases 1 and 2 (2). Still other models of repression propose that activator protein access to a DNA target sequence can be blocked by repressor protein binding to either the same site or an overlapping site. Such a mechanism appears to be operative in the competitive DNA binding of YY1 and serum response factor (SRF) 1 to serum response elements (SREs) of the c-fos promoter (5).An interesting variation on the theme of multiple proteins competing for overlapping binding sites arises from our attempts to elucidate the mechanism of vascular smooth muscle (VSM) ␣-actin pr...

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Section: Discussionmentioning

confidence: 99%

Cryptic MCAT Enhancer Regulation in Fibroblasts and Smooth Muscle Cells

Carlini

Getz

Strauch

et al. 2002

Journal of Biological Chemistry

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“…Stau1 to the Gag precursor (65)) or HIV-1 RNA (e.g. PABP1 binds to a region in the Gag coding sequence and controls Gag expression (70), whereas Pur␣, YB-1, and Stau1 bind to the TAR sequence (56,(71)(72)(73)). Other proteins of potential interest include hnRNP Q that associates with APOBEC1 (74) and interleukin-binding proteins 2 and 3 (NF45 and NF90, respectively), which are essential for encapsidation and protein priming of hepatitis B viral polymerase (75).…”

Section: Discussionmentioning

confidence: 99%

The Anti-HIV-1 Editing Enzyme APOBEC3G Binds HIV-1 RNA and Messenger RNAs That Shuttle between Polysomes and Stress Granules

Kozak

Marin

Rose

et al. 2006

Journal of Biological Chemistry

171

201

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Deoxycytidine deaminases APOBEC3G (A3G) and APOBEC3F (A3F) (members of the apolipoprotein B mRNA-editing catalytic polypeptide 3 family) have RNA-binding motifs, invade assembling human immunodeficiency virus (HIV-1), and hypermutate reverse transcripts. Antagonistically, HIV-1 viral infectivity factor degrades these enzymes. A3G is enzymatically inhibited by binding RNA within an unidentified large cytosolic ribonucleoprotein, implying that RNA degradation during reverse transcription may activate intravirion A3G at the necessary moment. We purified a biologically active tandem affinity-tagged A3G from human HEK293T cells. Mass spectrometry and coimmunoprecipitation from HEK293T and T lymphocyte extracts identified many RNAbinding proteins specifically associated with A3G and A3F, including poly(A)-binding proteins (PABPs), YB-1, Ro-La, RNA helicases, ribosomal proteins, and Staufen1. Most strikingly, nearly all A3G-associated proteins were known to bind exclusively or intermittently to translating and/or dormant mRNAs. Accordingly, A3G in HEK293T and T lymphocyte extracts was almost completely in A3G-mRNA-PABP complexes that shifted reversibly between polysomes and dormant pools in response to translational inhibitors. For example arsenite, which inhibits 5-cap-dependent translational initiation, shifted mRNA-A3G-PABP from polysomes into stress granules in a manner that was blocked and reversed by the elongation inhibitor cycloheximide. Immunofluorescence microscopy showed A3G-mRNA-PABP stress granules only partially overlapping with Staufen1. A3G coimmunoprecipitated HIV-1 RNA and many mRNAs. Ribonuclease released nearly all A3G-associated proteins, including A3G homo-oligomers and A3G-A3F hetero-oligomers, but the viral infectivity factor remained bound. Many proteins and RNAs associated with A3G are excluded from A3G-containing virions, implying that A3G competitively partitions into virions based on affinity for HIV-1 RNA.The viral infectivity factor (Vif) 4 encoded by human immunodeficiency virus type-1 (HIV-1) neutralizes a potent anti-HIV-1 defense system that occurs specifically in T lymphocytes and to a lesser degree in macrophages, thus explaining the efficient replication in T cells of wild-type HIV-1 but not HIV-1(⌬vif) that lacks a functional vif gene (1-4). This antiviral defense system involves the deoxycytidine deaminases APOBEC3G (A3G) and its paralog APOBEC3F (A3F) (members of the apolipoprotein B mRNA-editing catalytic polypeptide 3 family), each of which contains two RNA-binding motifs and incorporates into assembling HIV-1 capsids where they cause lethal dC-to-dU hypermutations in the single-stranded viral DNA that transiently forms during reverse transcription (1, 2, 4 -10). This single-stranded DNA, which has a minus sense, is synthesized using viral RNA as a template and is released from the DNA-RNA hybrid when the RNA is degraded by a ribonuclease H inherent in reverse transcriptase. Vif binds specifically to A3G and A3F and recruits a multisubunit ubiquitin ligase complex containing...

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“…27 For immunoprecipitation/Western blot analysis, approximately 300 mg of extracts (from cells producing C/EBPb, CFP-Tat, p27 SJ or its deletion mutants) were incubated with 2 ml of anti-C/EBPb (H-7), anti-Myc antibody (Invitrogen), polyclonal anti-p27 SJ , antibody specific for CFP-Tat (Living Colors, BD Biosciences Clontech) overnight at 41C. In all, 30 ml of Protein A Sepharose were added, the samples were incubated at 41C for 1 h, and resins were pelleted, washed, and immunoprecipitated bound complexes were resolved by SDS-PAGE and analyzed by Western blot utilizing antip27 SJ antibody (rabbit polyclonal antibody obtained from Lampire Biological Laboratories Inc., Pipersville, PA, USA).…”

Section: Protein-protein Interactionsmentioning

confidence: 99%