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Crystal structure of importin-? bound to the nuclear localization signal of Epstein-Barr virus EBNA-LP protein.

ABSTRACT: Epstein-Barr virus EBNA-LP protein is a transcriptional coactivator of EBNA2. Efficient nuclear localization of EBNA-LP is essential for cooperation with EBNA2. Here, we report the crystal structure of the nuclear import adaptor importin-?1 bound to the nuclear localization signal (NLS) of EBNA-LP that shows EBNA-LP residues 44-RRVRRR-49 binding to the major NLS-binding site at the P0-P5 positions. In contrast to previously characterized classical NLSs that invariably have a basic residue [either lysine (in the vast majority of cases) or arginine] at the P2 position, the EBNA-LP NLS is unique in that it has valine at the P2 position. The loss of the critical P2 lysine (or arginine) is compensated by arginine at the P0 position in the EBNA-LP NLS.

SUBMITTER: Nakada R

PROVIDER: S-EPMC5441407 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Crystal structure of importin-α bound to the nuclear localization signal of Epstein-Barr virus EBNA-LP protein.

Nakada Ryohei R Matsuura Yoshiyuki Y

Protein science : a publication of the Protein Society 20170417 6

Epstein-Barr virus EBNA-LP protein is a transcriptional coactivator of EBNA2. Efficient nuclear localization of EBNA-LP is essential for cooperation with EBNA2. Here, we report the crystal structure of the nuclear import adaptor importin-α1 bound to the nuclear localization signal (NLS) of EBNA-LP that shows EBNA-LP residues 44-RRVRRR-49 binding to the major NLS-binding site at the P0-P5 positions. In contrast to previously characterized classical NLSs that invariably have a basic residue [eithe ...[more]

PMID: 28383161

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Project description:Epstein-Barr Virus (EBV) is associated with numerous cancers including B cell lymphomas. In vitro, EBV transforms primary B cells into immortalized Lymphoblastoid Cell Lines (LCLs) which serves as a model to study the role of viral proteins in EBV malignancies. EBV induced cellular transformation is driven by viral proteins including EBV-Nuclear Antigens (EBNAs). EBNA-LP is important for the transformation of naïve but not memory B cells. While EBNA-LP was thought to promote gene activation by EBNA2, EBNA-LP Knock Out (LPKO) virus-infected cells express EBNA2-activated cellular genes efficiently. Therefore, a gap in knowledge exists as to what roles EBNA-LP plays in naïve B cell transformation. We developed a trans-complementation assay wherein transfection with wild-type EBNA-LP rescues the transformation of peripheral blood- and cord blood-derived naïve B cells by LPKO virus. Despite EBNA-LP phosphorylation sites being important in EBNA2 co-activation; neither phospho-mutant nor phospho-mimetic EBNA-LP was defective in rescuing naïve B cell outgrowth. However, we identified conserved leucine-rich motifs in EBNA-LP that were required for transformation of adult naïve and cord blood B cells. Because cellular PPAR-g coactivator (PGC) proteins use leucine-rich motifs to engage transcription factors including YY1, a key regulator of DNA looping and metabolism, we examined the role of EBNA-LP in engaging cellular transcription factors. We found a significant overlap between EBNA-LP and YY1 in ChIP-Seq data. By Cut&Run, YY1 peaks unique to WT compared to LPKO LCLs occur at more highly expressed genes. Moreover, Cas9 knockout of YY1 in primary B cells prior to EBV infection indicted YY1 to be essential for EBV-mediated transformation. We confirmed EBNA-LP and YY1and confirmed their biochemical association in LCLs by endogenous co-immunoprecipitation and. Moreover, we found that the EBNA-LP leucine-rich motifs were required for YY1 interaction in LCLs. Finally, we used Cas9 to knockout YY1 in primary total B cells and naïve B cells prior to EBV infection and found YY1 to be essential for EBV-mediated transformation. We propose that EBNA-LP engages YY1 through conserved leucine-rich motifs to promote EBV transformation of naïve B cells.

Dataset Information

Crystal structure of importin-? bound to the nuclear localization signal of Epstein-Barr virus EBNA-LP protein.

Publications

Crystal structure of importin-α bound to the nuclear localization signal of Epstein-Barr virus EBNA-LP protein.

Similar Datasets

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