Interferon Regulatory Factor 4 (IRF-4) Targets IRF-5 to Regulate Epstein-Barr Virus Transformation

The cellular interferon regulatory factor-4 (IRF-4), which is a member of IRF family, is involved in the development of multiple myeloma and Epstein-Barr virus (EBV)-mediated transformation of B lymphocytes. However, the molecular mechanism of IRF-4 in cellular transformation is unknown. We have found that knockdown of IRF-4 leads to high expression of IRF-5, a pro-apoptotic member in the IRF family. Overexpression of IRF-4 represses IRF-5 expression. Reduction of IRF-4 leads to growth inhibition, and the restoration of IRF-4 by exogenous plasmids correlates with the growth recovery and reduces IRF-5 expression. In addition, IRF-4 negatively regulates IRF-5 promoter reporter activities and binds to IRF-5 promoters in vivo and in vitro. Knockdown of IRF-5 rescues IRF-4 knockdownmediated growth inhibition, and IRF-5 overexpression alone is sufficient to induce cellular growth inhibition of EBV-transformed cells. Therefore, IRF-5 is one of the targets of IRF-4, and IRF-4 regulates the growth of EBV-transformed cells partially through IRF-5. This work provides insight on how IRFs interact with one another to participate in viral pathogenesis and transformation

Kaposi’s Sarcoma-Associated Herpesvirus Reduces Cellular Myeloid Differentiation Primary-Response Gene 88 (MyD88) Expression via Modulation of Its RNA

Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) is a human gammaherpesvirus associated with several human malignancies. The replication and transcription activator (RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication. Interleukin 1 (IL-1) is a major mediator for inflammation and plays an important role in both innate and adaptive immunity. Myeloid differentiation primary response gene 88 (MyD88) is an essential adaptor molecule for IL-1 as well as most Toll-like receptor signaling. In this study, we identified a novel mechanism by which KSHV interferes with host inflammation and immunity. KSHV RTA specifically reduces the steady-state protein levels of MyD88, and physiological levels of MyD88 are downregulated during KSHV lytic replication when RTA is expressed. The N-terminal region of RTA is required for the reduction of MyD88. Additional studies demonstrated that RTA targets MyD88 expression at the RNA level, inhibits RNA synthesis of MyD88, and may bind MyD88 RNA. Finally, RTA inhibits IL-1-mediated activation of NF-B. Because IL-1 is abundant in the KS microenvironment and inhibits KSHV replication, this work may expand our understanding of how KSHV evades host inflammation and immunity for its survival in vivo

Kaposi’s Sarcoma-Associated Herpesvirus Reduces Cellular Myeloid Differentiation Primary-Response Gene 88 (MyD88) Expression via Modulation of Its RNA

Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) is a human gammaherpesvirus associated with several human malignancies. The replication and transcription activator (RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication. Interleukin 1 (IL-1) is a major mediator for inflammation and plays an important role in both innate and adaptive immunity. Myeloid differentiation primary response gene 88 (MyD88) is an essential adaptor molecule for IL-1 as well as most Toll-like receptor signaling. In this study, we identified a novel mechanism by which KSHV interferes with host inflammation and immunity. KSHV RTA specifically reduces the steady-state protein levels of MyD88, and physiological levels of MyD88 are downregulated during KSHV lytic replication when RTA is expressed. The N-terminal region of RTA is required for the reduction of MyD88. Additional studies demonstrated that RTA targets MyD88 expression at the RNA level, inhibits RNA synthesis of MyD88, and may bind MyD88 RNA. Finally, RTA inhibits IL-1-mediated activation of NF-B. Because IL-1 is abundant in the KS microenvironment and inhibits KSHV replication, this work may expand our understanding of how KSHV evades host inflammation and immunity for its survival in vivo

Kaposi’s Sarcoma-Associated Herpesvirus Reduces Cellular Myeloid Differentiation Primary-Response Gene 88 (MyD88) Expression via Modulation of Its RNA

Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) is a human gammaherpesvirus associated with several human malignancies. The replication and transcription activator (RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication. Interleukin 1 (IL-1) is a major mediator for inflammation and plays an important role in both innate and adaptive immunity. Myeloid differentiation primary response gene 88 (MyD88) is an essential adaptor molecule for IL-1 as well as most Toll-like receptor signaling. In this study, we identified a novel mechanism by which KSHV interferes with host inflammation and immunity. KSHV RTA specifically reduces the steady-state protein levels of MyD88, and physiological levels of MyD88 are downregulated during KSHV lytic replication when RTA is expressed. The N-terminal region of RTA is required for the reduction of MyD88. Additional studies demonstrated that RTA targets MyD88 expression at the RNA level, inhibits RNA synthesis of MyD88, and may bind MyD88 RNA. Finally, RTA inhibits IL-1-mediated activation of NF-B. Because IL-1 is abundant in the KS microenvironment and inhibits KSHV replication, this work may expand our understanding of how KSHV evades host inflammation and immunity for its survival in vivo

TLR-TRIF Pathway Enhances the Expression of KSHV Replication and Transcription Activator

Background: Host innate immunity is against virus infection and replication. Results: Toll-like receptor 3 activation leads to enhanced expression of a key Kaposi’s sarcoma-associated herpesvirus (KSHV) protein. Conclusion: KSHV uses host Toll-like receptor pathway to augment its critical gene expression. Significance: A virus may usurp host innate immunity for its own benefits

Toll-Like Receptor 7 Stimulates the Expression of Epstein-Barr Virus Latent Membrane Protein 1

<div><p>Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus. Toll-like receptor 7 (TLR7) is involved in host innate immunity against pathogens, and its aberrant activation is linked to the development of systemic lupus erythematosus (SLE, also called “lupus”). Type I interferons (IFN) are apparently driving forces for lupus pathogenesis. Previously, we found that EBV latent membrane protein 1 (LMP1) primes cells for IFN production. In this report, the relationship among EBV LMP1, TLRs, and IFN production are examined. We find that TLR7 activation increases the expression of EBV LMP1, and IFN regulatory factor 7 (IRF7) is involved in the stimulation process. TLR7 activation did not induce IFNs from EBV-infected cells, but potentiates those cells for IFN production by TLR3 or TLR9 activation. In addition, we find that LMP1 and IFNs are co-expressed in the same cells in some lupus patients. Therefore, the aberrant activation of TLR7 might induce LMP1 expression and LMP1-expression cells may be producing IFNs in lupus patients. These results suggest EBV might be an exacerbating factor in some lupus patients via promoting IFN production.</p> </div

TLR7 activation stimulates the expression of EBV LMP1.

<p>A. TLR7 agonist stimulates EBV LMP1. SavIII or IB4 cells were treated with TLR7 agonist (imiquimod; 25 µg/ml), TLR3 agonist (poly (I:C); 10 µg/ml) or TLR9 agonist (ODN2395; 2.5 or 5 µM) for 24 hours. Cell lysates were used for Western blot analysis with LMP1 and GAPDH antibodies. The identity of proteins is as shown. B. TLR7 activation increases detectable LMP1-postive cells. IB4 cells were treated with imiquimod (25 µg/ml), for 24 hours, and the cells were fixed and stained with LMP1 primary and Alexa Fluor 647-labeled secondary antibodies. DAPI was used to stain the nuclei. The images were captured under identical conditions. The colors were artificially mounted to facilitate viewing. Blue, nuclei; red, LMP1. C. Quantification of detectable LMP1-positive cells. IB4 cells were treated and stained as in Panel B. The percentages of LMP1-positive cells were counted in 10 randomly selected fields. For untreated controls, the average number of cells per field with standard deviation is 201.5±59.5; while in imiquimod treated cells, the average number of cells per field with standard deviation is 170.6±70.8. The difference is statistically significant (<i>p</i><0.01). The <i>p</i> value was calculated by paired Student's t test with the use of Microsoft Excel.</p

EBV lytic replication plays limited role in the induction of LMP1.

<p>A. TLR7 activation induces LMP1 in EBV-transformed cells defective for viral lytic replication. An EBV-BRLF-null virus (RKO) transformed primary B cell line (LCL-RKO) and its parental virus transformed cells (LCL-wtEBV) were treated with imiquimod overnight. Cell lysates were used for Western blot analysis with LMP1 and tubulin antibodies. The identity of proteins is as shown. B. TLR7 activation failed to induce EBV lytic replication in EBV-transformed cells. LCL-wtEBV and LCL-RKO cells were treated with imiquimod overnight. The positive control was Akata cells treated with anti-human IgG. Cell lysates were used for Western blot analysis with LMP1 and Tubulin antibodies. The identity of proteins is as shown.</p