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Mutual inhibition between Kaposi's sarcoma-associated herpesvirus and Epstein-Barr virus lytic replication initiators in dually-infected primary effusion lymphoma.

Jiang Y, Xu D, Zhao Y, Zhang L - PLoS ONE (2008)

Bottom Line: Both Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are members of the human gamma herpesvirus family: each is associated with various human cancers.The majority of AIDS-associated primary effusion lymphoma (PEL) are co-infected with both KSHV and EBV.The leucine heptapeptide repeat (LR) region in K-RTA and leucine zipper region in EBV-Z are involved in the physical interactions of the two molecules.

View Article: PubMed Central - PubMed

Affiliation: Nebraska Center for Virology, University of Nebraska, Lincoln, Nebraska, USA.

ABSTRACT

Background: Both Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are members of the human gamma herpesvirus family: each is associated with various human cancers. The majority of AIDS-associated primary effusion lymphoma (PEL) are co-infected with both KSHV and EBV. Dually-infected PELs selectively switch from latency to lytic replication of either KSHV or EBV in response to chemical stimuli. KSHV replication and transcription activator (K-RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication, while EBV BZLF1 gene product (EBV-Z) is a critical initiator for induction of EBV lytic replication.

Methodology/principal findings: We show K-RTA and EBV-Z are co-localized and physically interact with each other in dually-infected PELs. K-RTA inhibits the EBV lytic replication by ifying EBV-Z-mediated EBV lytic gene activation. EBV-Z inhibits KSHV lytic gene expression by blocking K-RTA-mediated transactivations. The physical interaction between K-RTA and EBV-Z are required for the mutual inhibition of the two molecules. The leucine heptapeptide repeat (LR) region in K-RTA and leucine zipper region in EBV-Z are involved in the physical interactions of the two molecules. Finally, initiation of KSHV lytic gene expression is correlated with the reduction of EBV lytic gene expression in the same PEL cells.

Conclusions/significance: In this report, how the two viruses interact with each other in dually infected PELs is addressed. Our data may provide a possible mechanism for maintaining viral latency and for selective lytic replication in dually infected PELs, i.e., through mutual inhibition of two critical lytic replication initiators. Our data about putative interactions between EBV and KSHV would be applicable to the majority of AIDS-associated PELs and may be relevant to the pathogenesis of PELs.

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Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition.A. Schematic diagram of K-RTA domains and mutants. The DNA binding domain, leucine heptapeptide repeat region (LR), activation domain, and nuclear localization signal (NLS) are shown. The drawing is not on scale. In Panels B, C, and D, 293T cells were transfected with of the designated expression plasmids as shown on the top of the Figure. Cell extracts from these transfected cells were immunoprecipitated with either anti-EBV-Z (Panel B) or anti-K-RTA (Panel C). The immunoprecipitates were analyzed by Western blot (WB) using the indicated antibodies. In Panel D, whole cell lysate was used for western blot analyses. The identity of the respective proteins is denoted. E. Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition. 80 ng of K-RTA or K-RTA-DLR expression plasmids were transfected with various amounts of EBV-Z expression plasmid into BZLF1-KO (EBV+/KSHV−) cells as shown on the top. Lysates were used for western blot analysis 24 hours later. The same membrane was stripped and reprobed with other antibodies. The identity of proteins is as shown.
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pone-0001569-g004: Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition.A. Schematic diagram of K-RTA domains and mutants. The DNA binding domain, leucine heptapeptide repeat region (LR), activation domain, and nuclear localization signal (NLS) are shown. The drawing is not on scale. In Panels B, C, and D, 293T cells were transfected with of the designated expression plasmids as shown on the top of the Figure. Cell extracts from these transfected cells were immunoprecipitated with either anti-EBV-Z (Panel B) or anti-K-RTA (Panel C). The immunoprecipitates were analyzed by Western blot (WB) using the indicated antibodies. In Panel D, whole cell lysate was used for western blot analyses. The identity of the respective proteins is denoted. E. Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition. 80 ng of K-RTA or K-RTA-DLR expression plasmids were transfected with various amounts of EBV-Z expression plasmid into BZLF1-KO (EBV+/KSHV−) cells as shown on the top. Lysates were used for western blot analysis 24 hours later. The same membrane was stripped and reprobed with other antibodies. The identity of proteins is as shown.

Mentions: We suspect that K-RTA may physically interact with EBV-Z through its leucine heptapeptide repeat region (LR) of K-RTA [75]. This region is included within the domain of K-RTA required for interaction with the several cellular proteins such as K-RBP, RBP-Jκ, and C/EBPα [19], [23], [76], [77]. A mutant with the deletion of the region, K-RTA-DLR, was generated (Fig. 4A). The mutant protein is localized predominantly in the nucleus as wild type K-RTA (data not shown). The plasmids expressing EBV-Z, K-RTA and its mutant were transfected into 293T cells, and the interaction between EBV-Z and the mutant K-RTA was examined. While wt K-RTA interacted with EBV-Z properly, the K-RTA-DLR failed to interact with EBV-Z (Fig. 4B, and 4C). The expression of these proteins in 293T cells were similar (Fig. 4D). Thus, the LR region of K-RTA was involved in the physical interaction with EBV-Z.


Mutual inhibition between Kaposi's sarcoma-associated herpesvirus and Epstein-Barr virus lytic replication initiators in dually-infected primary effusion lymphoma.

Jiang Y, Xu D, Zhao Y, Zhang L - PLoS ONE (2008)

Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition.A. Schematic diagram of K-RTA domains and mutants. The DNA binding domain, leucine heptapeptide repeat region (LR), activation domain, and nuclear localization signal (NLS) are shown. The drawing is not on scale. In Panels B, C, and D, 293T cells were transfected with of the designated expression plasmids as shown on the top of the Figure. Cell extracts from these transfected cells were immunoprecipitated with either anti-EBV-Z (Panel B) or anti-K-RTA (Panel C). The immunoprecipitates were analyzed by Western blot (WB) using the indicated antibodies. In Panel D, whole cell lysate was used for western blot analyses. The identity of the respective proteins is denoted. E. Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition. 80 ng of K-RTA or K-RTA-DLR expression plasmids were transfected with various amounts of EBV-Z expression plasmid into BZLF1-KO (EBV+/KSHV−) cells as shown on the top. Lysates were used for western blot analysis 24 hours later. The same membrane was stripped and reprobed with other antibodies. The identity of proteins is as shown.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2215330&req=5

pone-0001569-g004: Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition.A. Schematic diagram of K-RTA domains and mutants. The DNA binding domain, leucine heptapeptide repeat region (LR), activation domain, and nuclear localization signal (NLS) are shown. The drawing is not on scale. In Panels B, C, and D, 293T cells were transfected with of the designated expression plasmids as shown on the top of the Figure. Cell extracts from these transfected cells were immunoprecipitated with either anti-EBV-Z (Panel B) or anti-K-RTA (Panel C). The immunoprecipitates were analyzed by Western blot (WB) using the indicated antibodies. In Panel D, whole cell lysate was used for western blot analyses. The identity of the respective proteins is denoted. E. Interaction between K-RTA and EBV-Z is required for K-RTA-mediated inhibition. 80 ng of K-RTA or K-RTA-DLR expression plasmids were transfected with various amounts of EBV-Z expression plasmid into BZLF1-KO (EBV+/KSHV−) cells as shown on the top. Lysates were used for western blot analysis 24 hours later. The same membrane was stripped and reprobed with other antibodies. The identity of proteins is as shown.
Mentions: We suspect that K-RTA may physically interact with EBV-Z through its leucine heptapeptide repeat region (LR) of K-RTA [75]. This region is included within the domain of K-RTA required for interaction with the several cellular proteins such as K-RBP, RBP-Jκ, and C/EBPα [19], [23], [76], [77]. A mutant with the deletion of the region, K-RTA-DLR, was generated (Fig. 4A). The mutant protein is localized predominantly in the nucleus as wild type K-RTA (data not shown). The plasmids expressing EBV-Z, K-RTA and its mutant were transfected into 293T cells, and the interaction between EBV-Z and the mutant K-RTA was examined. While wt K-RTA interacted with EBV-Z properly, the K-RTA-DLR failed to interact with EBV-Z (Fig. 4B, and 4C). The expression of these proteins in 293T cells were similar (Fig. 4D). Thus, the LR region of K-RTA was involved in the physical interaction with EBV-Z.

Bottom Line: Both Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are members of the human gamma herpesvirus family: each is associated with various human cancers.The majority of AIDS-associated primary effusion lymphoma (PEL) are co-infected with both KSHV and EBV.The leucine heptapeptide repeat (LR) region in K-RTA and leucine zipper region in EBV-Z are involved in the physical interactions of the two molecules.

View Article: PubMed Central - PubMed

Affiliation: Nebraska Center for Virology, University of Nebraska, Lincoln, Nebraska, USA.

ABSTRACT

Background: Both Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) are members of the human gamma herpesvirus family: each is associated with various human cancers. The majority of AIDS-associated primary effusion lymphoma (PEL) are co-infected with both KSHV and EBV. Dually-infected PELs selectively switch from latency to lytic replication of either KSHV or EBV in response to chemical stimuli. KSHV replication and transcription activator (K-RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication, while EBV BZLF1 gene product (EBV-Z) is a critical initiator for induction of EBV lytic replication.

Methodology/principal findings: We show K-RTA and EBV-Z are co-localized and physically interact with each other in dually-infected PELs. K-RTA inhibits the EBV lytic replication by ifying EBV-Z-mediated EBV lytic gene activation. EBV-Z inhibits KSHV lytic gene expression by blocking K-RTA-mediated transactivations. The physical interaction between K-RTA and EBV-Z are required for the mutual inhibition of the two molecules. The leucine heptapeptide repeat (LR) region in K-RTA and leucine zipper region in EBV-Z are involved in the physical interactions of the two molecules. Finally, initiation of KSHV lytic gene expression is correlated with the reduction of EBV lytic gene expression in the same PEL cells.

Conclusions/significance: In this report, how the two viruses interact with each other in dually infected PELs is addressed. Our data may provide a possible mechanism for maintaining viral latency and for selective lytic replication in dually infected PELs, i.e., through mutual inhibition of two critical lytic replication initiators. Our data about putative interactions between EBV and KSHV would be applicable to the majority of AIDS-associated PELs and may be relevant to the pathogenesis of PELs.

Show MeSH
Related in: MedlinePlus