A Locus Encompassing the Epstein-Barr Virus bglf4 Kinase Regulates Expression of Genes Encoding Viral Structural Proteins

El-Guindy, Ayman; López-Giráldez, Francesc; Delecluse, Henri Jacques; McKenzie, Jessica; Miller, George

doi:10.1371/journal.ppat.1004307

Cited by 33 publications

(36 citation statements)

References 73 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Indeed, ORF36-dependent phosphorylation of viral K-bzip has been shown to modulate viral transcription (16). Furthermore, EBV BGLF4 has been shown to play a critical role in the efficient expression of several EBV late lytic genes (36). (ii) ORF36 is critical for efficient viral DNA synthesis.…”

Section: Discussionmentioning

confidence: 99%

Discovery of a Coregulatory Interaction between Kaposi's Sarcoma-Associated Herpesvirus ORF45 and the Viral Protein Kinase ORF36

Avey

Tepper

Pifer

et al. 2016

J Virol

View full text Add to dashboard Cite

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of three human malignancies. KSHV ORF36 encodes a serine/threonine viral protein kinase, which is conserved throughout all herpesviruses. Although several studies have identified the viral and cellular substrates of conserved herpesvirus protein kinases (CHPKs), the precise functions of KSHV ORF36 during lytic replication remain elusive. Here, we report that ORF36 interacts with another lytic protein, ORF45, in a manner dependent on ORF36 kinase activity. We mapped the regions of ORF36 and ORF45 involved in the binding. Their association appears to be mediated by electrostatic interactions, since deletion of either the highly basic N terminus of ORF36 or an acidic patch of ORF45 abolished the binding. In addition, the dephosphorylation of ORF45 protein dramatically reduced its association with ORF36. Importantly, ORF45 enhances both the stability and kinase activity of ORF36. Consistent with previous studies of CHPK homologs, we detected ORF36 protein in extracellular virions. To investigate the roles of ORF36 in the context of KSHV lytic replication, we used bacterial artificial chromosome mutagenesis to engineer both ORF36-null and kinase-dead mutants. We found that ORF36-null/mutant virions are moderately defective in viral particle production and are further deficient in primary infection. In summary, our results uncover a functionally important interaction between ORF36 and ORF45 and indicate a significant role of ORF36 in the production of infectious progeny virions. IMPORTANCEKaposi's sarcoma-associated herpesvirus (KSHV) is a human tumor virus with a significant public health burden. KSHV ORF36 encodes a serine/threonine viral protein kinase, whose functions throughout the viral life cycle have not been elucidated. Here, we report that ORF36 interacts with another KSHV protein, ORF45. We mapped the regions of ORF36 and ORF45 involved in their association and further characterized the consequences of this interaction. We engineered ORF36 mutant viruses in order to investigate the functional roles of ORF36 in the context of KSHV lytic replication, and we confirmed that ORF36 is a component of KSHV virions. Moreover, we found that ORF36 mutants are defective in virion production and primary infection. In summary, we discovered and characterized a functionally important interaction between KSHV ORF36 and ORF45, and our results suggest a significant role of ORF36 in the production of infectious progeny virions, a process critical for KSHV pathogenesis. K aposi's sarcoma-associated herpesvirus (KSHV) is a human tumor virus and the causative agent of Kaposi's sarcoma (KS), as well as two lymphoproliferative disorders (1-3). All herpesviruses encode at least one serine/threonine protein kinase that is conserved throughout the three subfamilies (alpha-, beta-, and gammaherpesviruses), collectively referred to as conserved herpesvirus protein kinases (CHPKs) (reviewed in references 4 and 5). Orthologs of CHPKs include UL13 of herpes simp...

show abstract

Section: Discussionmentioning

confidence: 99%

Discovery of a Coregulatory Interaction between Kaposi's Sarcoma-Associated Herpesvirus ORF45 and the Viral Protein Kinase ORF36

Avey

Tepper

Pifer

et al. 2016

J Virol

View full text Add to dashboard Cite

show abstract

“…In the study by Reference , pBGLF3 was localized both in the nucleus and in the cytoplasm which is in line with our data. To our knowledge, the functional data available for pBGLF3 is limited to a report suggesting a role in late viral gene regulation . It remains to be seen how nucleo‐cytoplasmic shuttling contributes to this protein's function.…”

Section: Discussionmentioning

confidence: 99%

“…To our knowledge, the functional data available for pBGLF3 is limited to a report suggesting a role in late viral gene regulation. 40 It remains to be seen how nucleo-cytoplasmic shuttling contributes to this protein's function. The second exported EBV protein, pBTRF1 and its HSV1 ortholog pUL21 were localized in both nucleus and cytoplasm, 5 and both were found to be actively exported in the NEX-TRAP assay.…”

Section: Analysis Of This Large Group Of Viral Tegument Proteins Demon-mentioning

confidence: 99%

Comprehensive analysis of nuclear export of herpes simplex virus type 1 tegument proteins and their Epstein‐Barr virus orthologs

Funk

Raschbichler

Lieber

et al. 2019

Traffic

View full text Add to dashboard Cite

Morphogenesis of herpesviral virions is initiated in the nucleus but completed in the cytoplasm. Mature virions contain more than 25 tegument proteins many of which perform both nuclear and cytoplasmic functions suggesting they shuttle between these compartments. While nuclear import of herpesviral proteins was shown to be crucial for viral propagation, active nuclear export and its functional impact are still poorly understood. To systematically analyze nuclear export of tegument proteins present in virions of Herpes simplex virus type 1 (HSV1) and Epstein‐Barr virus (EBV), the Nuclear EXport Trapped by RAPamycin (NEX‐TRAP) was applied. Nine of the 22 investigated HSV1 tegument proteins including pUL4, pUL7, pUL11, pUL13, pUL21, pUL37d11, pUL47, pUL48 and pUS2 as well as 2 out of 6 EBV orthologs harbor nuclear export activity. A functional leucine‐rich nuclear export sequence (NES) recognized by the export factor CRM1/Xpo1 was identified in six of them. The comparison between experimental and bioinformatic data indicates that experimental validation of predicted NESs is required. Mutational analysis of the pUL48/VP16 NES revealed its importance for herpesviral propagation. Together our data suggest that nuclear export is an important feature of the herpesviral life cycle required to co‐ordinate nuclear and cytoplasmic processes.

show abstract

“…The DNA was purified with a Qiagen PCR purification kit. Viral genome amplification was measured using iQ SY3R green supermix (Bio-Rad) with primers targeting oriLyt (34). See Table S1 in the supplemental material for primer sequences.…”

Section: Methodsmentioning

confidence: 99%

A Noncanonical Basic Motif of Epstein-Barr Virus ZEBRA Protein Facilitates Recognition of Methylated DNA, High-Affinity DNA Binding, and Lytic Activation

Weber

Buzovetsky

Heston

et al. 2019

J Virol

Self Cite

View full text Add to dashboard Cite

The pathogenesis of Epstein-Barr virus (EBV) infection, including development of lymphomas and carcinomas, is dependent on the ability of the virus to transit from latency to the lytic phase. This conversion, and ultimately disease development, depends on the molecular switch protein, ZEBRA, a viral bZIP transcription factor that initiates transcription from promoters of viral lytic genes. By binding to the origin of viral replication, ZEBRA is also an essential replication protein. Here, we identified a novel DNA-binding motif of ZEBRA, N terminal to the canonical bZIP domain. This RRTRK motif is important for high-affinity binding to DNA and is essential for recognizing the methylation state of viral promoters. Mutations in this motif lead to deficiencies in DNA binding, recognition of DNA methylation, lytic cycle DNA replication, and viral late gene expression. This work advances our understanding of ZEBRA-dependent activation of the viral lytic cascade. IMPORTANCE The binding of ZEBRA to methylated and unmethylated viral DNA triggers activation of the EBV lytic cycle, leading to viral replication and, in some patients, cancer development. Our work thoroughly examines how ZEBRA uses a previously unrecognized basic motif to bind nonmethylated and methylated DNA targets, leading to viral lytic activation. Our findings show that two different positively charged motifs, including the canonical BZIP domain and a newly identified RRTRK motif, contribute to the mechanism of DNA recognition by a viral AP-1 protein. This work contributes to the assessment of ZEBRA as a potential therapeutic target for antiviral and oncolytic treatments.

show abstract

A Locus Encompassing the Epstein-Barr Virus bglf4 Kinase Regulates Expression of Genes Encoding Viral Structural Proteins

Cited by 33 publications

References 73 publications

Discovery of a Coregulatory Interaction between Kaposi's Sarcoma-Associated Herpesvirus ORF45 and the Viral Protein Kinase ORF36

Discovery of a Coregulatory Interaction between Kaposi's Sarcoma-Associated Herpesvirus ORF45 and the Viral Protein Kinase ORF36

Comprehensive analysis of nuclear export of herpes simplex virus type 1 tegument proteins and their Epstein‐Barr virus orthologs

A Noncanonical Basic Motif of Epstein-Barr Virus ZEBRA Protein Facilitates Recognition of Methylated DNA, High-Affinity DNA Binding, and Lytic Activation

Contact Info

Product

Resources

About