Dataset Information

Binding of RBP-Jkappa (CSL) protein to the promoter of the Kaposi's sarcoma-associated herpesvirus ORF47 (gL) gene is a critical but not sufficient determinant of transactivation by ORF50 protein.

ABSTRACT: ORF50 protein activates the KSHV lytic cycle. The promoter of an early lytic-cycle gene ORF47, encoding envelope protein gL, is activated by an interaction between ORF50 protein and RBP-Jkappa. In ORF47p only one of two sequences fitting the consensus RBP-Jkappa recognition site strongly binds RBP-Jkappa and confers a response to ORF50 protein. Flanking sequences 5' to the RBP-Jkappa binding site are required to confer a maximal response to ORF50 protein. Not all mutant ORF50 response elements in the ORF47p that are bound by RBP-Jkappa are sufficient to confer maximal ORF50 responsiveness. Four sequences containing an RBP-Jkappa site and flanking sequences characteristic of the ORF50 response element in ORF47p were identified in human DNA. All bound RBP-Jkappa, but only one responded robustly to ORF50 protein. We propose models for the possible function of ancillary sequences flanking the RBP-Jkappa-binding element which are crucial for mediating ORF50 transactivation.

SUBMITTER: Chang PJ

PROVIDER: S-EPMC2823930 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Binding of RBP-Jkappa (CSL) protein to the promoter of the Kaposi's sarcoma-associated herpesvirus ORF47 (gL) gene is a critical but not sufficient determinant of transactivation by ORF50 protein.

Chang Pey-Jium PJ Boonsiri Joseph J Wang Shih-Shan SS Chen Li-Yu LY Miller George G

Virology 20091216 1

ORF50 protein activates the KSHV lytic cycle. The promoter of an early lytic-cycle gene ORF47, encoding envelope protein gL, is activated by an interaction between ORF50 protein and RBP-Jkappa. In ORF47p only one of two sequences fitting the consensus RBP-Jkappa recognition site strongly binds RBP-Jkappa and confers a response to ORF50 protein. Flanking sequences 5' to the RBP-Jkappa binding site are required to confer a maximal response to ORF50 protein. Not all mutant ORF50 response elements i ...[more]

PMID: 20006367

Similar Datasets

Project description:The Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded open reading frame 50 (ORF50) protein is the key transactivator responsible for the latent-to-lytic switch. Here, we investigated the transcriptional activation of the ORF56 gene (encoding a primase protein) by ORF50 and successfully identified an ORF50-responsive element located in the promoter region between positions -97 and -44 (designated 56p-RE). This 56p-RE element contains a noncanonical RBP-Jκ-binding sequence and a nonconsensus Sp1/Sp3-binding sequence. Electrophoretic mobility shift assays revealed that RBP-Jκ, Sp3, and ORF50 could form stable complexes on the 56p-RE element. Importantly, transient-reporter analysis showed that Sp3, but not RBP-Jκ or Sp1, acts in synergy with ORF50 to activate the 56p-RE-containing reporter construct, and the synergy mainly depends on the Sp1/Sp3-binding region of the 56p-RE element. Sequence similarity searches revealed that the promoters for ORF21 (thymidine kinase), ORF60 (ribonucleotide reductase, small subunit), and cellular interleukin-10 (IL-10) contain a sequence motif similar to the Sp1/Sp3-binding region of the 56p-RE element, and we found that these promoters could also be synergistically activated by ORF50 and Sp3 via the conserved motifs. Noteworthily, the conversion of the Sp1/Sp3-binding sequence of the 56p-RE element into a consensus high-affinity Sp-binding sequence completely lost the synergistic response to ORF50 and Sp3. Moreover, transcriptional synergy could not be detected through other ORF50-responsive elements from the viral PAN, K12, ORF57, and K6 promoters. Collectively, the results of our study demonstrate that ORF50 and Sp3 can act in synergy on the transcription of specific gene promoters, and we find a novel conserved cis-acting motif in these promoters essential for transcriptional synergy.IMPORTANCE Despite the critical role of ORF50 in the KSHV latent-to-lytic switch, the molecular mechanism by which ORF50 activates its downstream target genes, especially those that encode the viral DNA replication enzymes, is not yet fully understood. Here, we find that ORF50 can cooperate with Sp3 to synergistically activate promoters of the viral ORF56 (primase), ORF21 (thymidine kinase), and ORF60 (ribonucleotide reductase) genes via similar Sp1/Sp3-binding motifs. Additionally, the same synergistic effect can be seen on the promoter of the cellular IL-10 gene. Overall, our data reveal an important role for Sp3 in ORF50-mediated transactivation, and we propose a new subclass of ORF50-responsive elements in viral and cellular promoters.

Dataset Information

Binding of RBP-Jkappa (CSL) protein to the promoter of the Kaposi's sarcoma-associated herpesvirus ORF47 (gL) gene is a critical but not sufficient determinant of transactivation by ORF50 protein.

Publications

Binding of RBP-Jkappa (CSL) protein to the promoter of the Kaposi's sarcoma-associated herpesvirus ORF47 (gL) gene is a critical but not sufficient determinant of transactivation by ORF50 protein.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure