Kaposi's Sarcoma-Associated Herpesvirus Rta Tetramers Make High-Affinity Interactions with Repetitive DNA Elements in the Mta Promoter To Stimulate DNA Binding of RBP-Jk/CSL

Palmeri, Diana; Carroll, Kyla Driscoll; González‐López, Olga; Lukac, David M.

doi:10.1128/jvi.05479-11

Cited by 20 publications

(81 citation statements)

References 52 publications

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“…pcDNA3, pcDNA3-FLg50, and pcDNA-FLc50 were previously described (16,(31)(32)(33). HDAC1-FLAG, HDAC3-FLAG, HDAC6-FLAG, HDAC7-FLAG, and SIRT1-FLAG (34-36) (gifts of Eric Verdin) and H2b-green fluorescent protein (GFP) (37) (a gift of Geoff Wahl) were obtained from the AddGene repository (plasmids 13820, 13819, 13823, 13824, 13812, and 11680, respectively).…”

Section: Methodsmentioning

confidence: 99%

Histone Deacetylase Classes I and II Regulate Kaposi's Sarcoma-Associated Herpesvirus Reactivation

Shin

DeCotiis

Giron

et al. 2014

J Virol

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In primary effusion lymphoma (PEL) cells infected with latent Kaposi's sarcoma-associated herpesvirus (KSHV), the promoter of the viral lytic switch gene, Rta, is organized into bivalent chromatin, similar to cellular developmental switch genes. Histone deacetylase (HDAC) inhibitors (HDACis) reactivate latent KSHV and dramatically remodel the viral genome topology and chromatin architecture. However, reactivation is not uniform across a population of infected cells. We sought to identify an HDACi cocktail that would uniformly reactivate KSHV and reveal the regulatory HDACs. Using HDACis with various specificities, we found that class I HDACis were sufficient to reactivate the virus but differed in potency. Valproic acid (VPA) was the most effective HDACi, inducing lytic cycle gene expression in 75% of cells, while trichostatin A (TSA) induced less widespread lytic gene expression and inhibited VPA-stimulated reactivation. VPA was only slightly superior to TSA in inducing histone acetylation of Rta's promoter, but only VPA induced significant production of infectious virus, suggesting that HDAC regulation after Rta expression has a dramatic effect on reactivation progression. Ectopic HDACs 1, 3, and 6 inhibited TPA-stimulated KSHV reactivation. Surprisingly, ectopic HDACs 1 and 6 stimulated reactivation independently, suggesting that the stoichiometries of HDAC complexes are critical for the switch. Tubacin, a specific inhibitor of the ubiquitin-binding, proautophagic HDAC6, also inhibited VPA-stimulated reactivation. Immunofluorescence indicated that HDAC6 is expressed diffusely throughout latently infected cells, but its expression level and nuclear localization is increased during reactivation. Overall, our data suggest that inhibition of HDAC classes I and IIa and maintenance of HDAC6 (IIb) activity are required for optimal KSHV reactivation.

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Section: Methodsmentioning

confidence: 99%

Histone Deacetylase Classes I and II Regulate Kaposi's Sarcoma-Associated Herpesvirus Reactivation

Shin

DeCotiis

Giron

et al. 2014

J Virol

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“…A doxycycline (Dox)-inducible Rta cell line TREx BCBL-1-Rta (a gift of J. Jung, University of Southern California [USC] [38]), a wild-type, KSHV-infected B lymphoma cell line BCBL-1, and uninfected B lymphoma cell line BL-41 were maintained in RPMI 1640 supplemented with 12% fetal bovine serum (Sigma) as previously described (15,22 Vieira), were maintained as previously described (15,22,(39)(40)(41)(42)(43)(44). iSLK cell lines contain Dox-inducible Rta and were gifts of D. Ganem (42).…”

Section: Methodsmentioning

confidence: 99%

“…For luciferase reporter assays, up to 27 g total DNA was used per transfection. pcDNA3.1-His-lacZ was cotransfected as an internal control, and normalization with ␤-galactosidase was performed as previously described (22). For viral antigen expression, at least 700 cells per technical replicate were counted.…”

Section: Methodsmentioning

confidence: 99%

“…All plasmids were amplified, purified, and sequenced as previously described (22). pcDNA3 is the empty vector control, and pcDNA3.1-His-lacZ, expresses His-tagged ␤-galactosidase (Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

“…Though Rta expression is sufficient to reactivate KSHV in a population of cells, single-cell assays suggest that it is not sufficient to reactivate the virus uniformly in every Rta-expressing cell (13,15). Rta, a 120-kDa transcription factor, directly transactivates downstream viral and cellular genes through interactions with essential cofactors, including KSHV delayed early protein Mta (ORF57) (15)(16)(17)(18) and cellular Notch pathway effector recombination-signal binding protein (RBP-Jk) (19)(20)(21)(22).…”

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confidence: 99%

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The Cellular Peptidyl-Prolyl cis / trans Isomerase Pin1 Regulates Reactivation of Kaposi's Sarcoma-Associated Herpesvirus from Latency

Guito

Gavina

Palmeri

et al. 2014

J Virol

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Kaposi's sarcoma-associated herpesvirus (KSHV) causes Kaposi's sarcoma and primary effusion lymphoma. KSHV-infected cells are predominantly latent, with a subset undergoing lytic reactivation. Rta is the essential lytic switch protein that reactivates virus by forming transactivation-competent complexes with the Notch effector protein RBP-Jk and promoter DNA. Strikingly, Rta homolog analysis reveals that prolines constitute 17% of conserved residues. Rta is also highly phosphorylated in vivo. We previously demonstrated that proline content determines Rta homotetramerization and function. We hypothesize that prolinedirected modifications regulate Rta function by controlling binding to peptidyl-prolyl cis/trans isomerases (PPIases). Cellular PPIase Pin1 binds specifically to phosphoserine-or phosphothreonine-proline (pS/T-P) motifs in target proteins. Pin1 dysregulation is implicated in myriad human cancers and can be subverted by viruses. Our data show that KSHV Rta protein contains potential pS/T-P motifs and binds directly to Pin1. Rta transactivation is enhanced by Pin1 at two delayed early viral promoters in uninfected cells. Pin1's effect, however, suggests a rheostat-like influence on Rta function. We show that in infected cells, endogenous Pin1 is active during reactivation and enhances Rta-dependent early protein expression induced by multiple signals, as well as DNA replication. Surprisingly, ablation of Pin1 activity by the chemical juglone or dominant-negative Pin1 enhanced late gene expression and production of infectious virus, while ectopic Pin1 showed inhibitory effects. Our data thus suggest that Pin1 is a unique, dose-dependent molecular timer that enhances Rta protein function, but inhibits late gene synthesis and virion production, during KSHV lytic reactivation.

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Cancers in People with HIV and AIDS

Yarchoan¹

2014

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Kaposi's Sarcoma-Associated Herpesvirus Rta Tetramers Make High-Affinity Interactions with Repetitive DNA Elements in the Mta Promoter To Stimulate DNA Binding of RBP-Jk/CSL

Cited by 20 publications

References 52 publications

Histone Deacetylase Classes I and II Regulate Kaposi's Sarcoma-Associated Herpesvirus Reactivation

Histone Deacetylase Classes I and II Regulate Kaposi's Sarcoma-Associated Herpesvirus Reactivation

The Cellular Peptidyl-Prolyl cis / trans Isomerase Pin1 Regulates Reactivation of Kaposi's Sarcoma-Associated Herpesvirus from Latency

Cancers in People with HIV and AIDS

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