KSHV viral cyclin inactivates p27KIP1 through Ser10 and Thr187 phosphorylation in proliferating primary effusion lymphomas

Sarek, Grzegorz; Järviluoma, Annika; Ojala, Päivi M.

doi:10.1182/blood-2005-06-2534

Cited by 54 publications

(59 citation statements)

References 49 publications

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“…BC-3 cell lysates were separated by gel filtration chromatography on a Superdex 200 HR column with a fast protein liquid chromatography (FPLC) system (Pharmacia Biotech, Uppsala, Sweden) as described previously (38).…”

Section: Methodsmentioning

confidence: 99%

Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions

Paudel

Sadagopan

Chakraborty

et al. 2012

J Virol

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Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically associated with the angioproliferative Kaposi's sarcoma (KS). KSHV infection and the expression of latency-associated nuclear antigen (LANA-1) upregulates the angiogenic multifunctional 123-amino-acid, 14-kDa protein angiogenin (ANG), which is detected in KS lesions and in KSHV-associated primary effusion lymphoma (PEL) cells. ANG knockdown or the inhibition of ANG's nuclear translocation resulted in decreased LANA-1 gene expression and reduced KSHV-infected endothelial and PEL cell survival (Sadagopan et al., J. Virol. 83: 3342–3364, 2009). Further studies here demonstrate that LANA-1 and ANG colocalize and coimmunoprecipitate in de novo infected endothelial cells and in latently infected PEL (BCBL-1 and BC-3) cells. LANA-1 and ANG interaction occurred in the absence of the KSHV genome and other viral proteins. In gel filtration chromatography analyses of BC-3 cell lysates, ANG coeluted with LANA-1, p53, and Mdm2 in high-molecular-weight fractions, and LANA-1, p53, and Mdm2 also coimmunoprecipitated with ANG. LANA-1, ANG, and p53 colocalized in KSHV-infected cells, and colocalization between ANG and p53 was also observed in LANA-1-negative cells. The deletion constructs of ANG suggested that the C-terminal region of amino acids 104 to 123 is involved in LANA-1 and p53 interactions. Silencing ANG or inhibiting its nuclear translocation resulted in decreased nuclear LANA-1 and ANG levels, decreased interactions between ANG-LANA-1, ANG-p53, and LANA-1-p53, the induction of p53, p21, and Bax proteins, the increased cytoplasmic localization of p53, the downregulation of Bcl-2, the increased cleavage of caspase-3, and the apoptosis of cells. No such effects were observed in KSHV-negative BJAB cells. The phosphorylation of p53 at serine 15, which is essential for p53 stabilization and for p53's apoptotic and cell cycle regulation functions, was increased in BCBL-1 cells transduced with short hairpin RNA targeting ANG. Together, these studies suggest that the antiapoptosis observed in KSHV-infected cells and the suppression of p53 functions are mediated in part by ANG, and KSHV has probably evolved to utilize angiogenin's multiple functions for the maintenance of its latency and cell survival. Thus, targeting ANG to induce the apoptosis of cells latently infected with KSHV is an attractive therapeutic strategy against KSHV infection and associated malignancies.

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Section: Methodsmentioning

confidence: 99%

Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions

Paudel

Sadagopan

Chakraborty

et al. 2012

J Virol

Self Cite

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“…Western blotting analysis, immunoprecipitations, and gel filtration chromatography were carried out as described previously (51). Gel filtration chromatography was performed on a Superdex 200 HR column (Amersham Pharmacia).…”

Section: Cell Lines Pel Cell Line Bc-1 Is Derived From a Hiv-positivmentioning

confidence: 99%

“…HRP-conjugated antibodies specific for rabbit, mouse, or rat immunoglobulins were purchased from Chemicon International. Cytocentrifugation and indirect immunofluorescence were performed as previously described (51). PEL cell lines and EBV-transformed LCLs were labeled with a mouse monoclonal antibody against γH2AX (Upstate USA Inc.) and Alexa Fluor 594-conjugated antibody to rabbit immunoglobulin (Invitrogen).…”

Section: Cell Lines Pel Cell Line Bc-1 Is Derived From a Hiv-positivmentioning

confidence: 99%

Reactivation of the p53 pathway as a treatment modality for KSHV-induced lymphomas

Sarek¹,

Kurki²,

Enbäck³

et al. 2007

J. Clin. Invest.

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133

135

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Kaposi's sarcoma herpesvirus (KSHV) is the etiologic agent for primary effusion lymphoma (PEL), a nonHodgkin type lymphoma manifesting as an effusion malignancy in the affected individual. Although KSHV has been recognized as a tumor virus for over a decade, the pathways for its tumorigenic conversion are incompletely understood, which has greatly hampered the development of efficient therapies for KSHV-induced malignancies like PEL and Kaposi's sarcoma. There are no current therapies effective against the aggressive, KSHV-induced PEL. Here we demonstrate that activation of the p53 pathway using murine double minute 2 (MDM2) inhibitor Nutlin-3a conveyed specific and highly potent activation of PEL cell killing. Our results demonstrated that the KSHV latency-associated nuclear antigen (LANA) bound to both p53 and MDM2 and that the MDM2 inhibitor Nutlin-3a disrupted the p53-MDM2-LANA complex and selectively induced massive apoptosis in PEL cells. Together with our results indicating that KSHV-infection activated DNA damage signaling, these findings contribute to the specificity of the cytotoxic effects of Nutlin-3a in KSHV-infected cells. Moreover, we showed that Nutlin-3a had striking antitumor activity in vivo in a mouse xenograft model. Our results therefore present new options for exploiting reactivation of p53 as what we believe to be a novel and highly selective treatment modality for this virally induced lymphoma.

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“…Cell lysates obtained from BC-3 cells were separated by gel filtration chromatography on a Superdex 200 HR column with a fast protein liquid chromatography (FPLC) system (Pharmacia Biotech, Uppsala, Sweden) as described previously (39).…”

Section: Methodsmentioning

confidence: 99%

Activated Nrf2 Interacts with Kaposi's Sarcoma-Associated Herpesvirus Latency Protein LANA-1 and Host Protein KAP1 To Mediate Global Lytic Gene Repression

Gjyshi

Roy

Dutta

et al. 2015

J Virol

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Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically associated with Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease. We have previously shown that KSHV utilizes the host transcription factor Nrf2 to aid in infection of endothelial cells and oncogenesis. Here, we investigate the role of Nrf2 in PEL and PEL-derived cell lines and show that KSHV latency induces Nrf2 protein levels and transcriptional activity through the COX-2/PGE2/EP4/PKC axis. Next-generation sequencing of KSHV transcripts in the PEL-derived BCBL-1 cell line revealed that knockdown of this activated Nrf2 results in global elevation of lytic genes. Nrf2 inhibition by the chemical brusatol also induces lytic gene expression. Both Nrf2 knockdown and brusatol-mediated inhibition induced KSHV lytic reactivation in BCBL-1 cells. In a series of follow-up experiments, we characterized the mechanism of Nrf2-mediated regulation of KSHV lytic repression during latency. Biochemical assays showed that Nrf2 interacted with KSHV latency-associated nuclear antigen 1 (LANA-1) and the host transcriptional repressor KAP1, which together have been shown to repress lytic gene expression. Promoter studies showed that although Nrf2 alone induces the open reading frame 50 (ORF50) promoter, its association with LANA-1 and KAP1 abrogates this effect. Interestingly, LANA-1 is crucial for efficient KAP1/Nrf2 association, while Nrf2 is essential for LANA-1 and KAP1 recruitment to the ORF50 promoter and its repression. Overall, these results suggest that activated Nrf2, LANA-1, and KAP1 assemble on the ORF50 promoter in a temporal fashion. Initially, Nrf2 binds to and activates the ORF50 promoter during early de novo infection, an effect that is exploited during latency by LANA-1-mediated recruitment of the host transcriptional repressor KAP1 on Nrf2. Cell death assays further showed that Nrf2 and KAP1 knockdown induce significant cell death in PEL cell lines. Our studies suggest that Nrf2 modulation through available oral agents is a promising therapeutic approach in the treatment of KSHV-associated malignancies. IMPORTANCEKS and PEL are aggressive KSHV-associated malignancies with moderately effective, highly toxic chemotherapies. Other than ganciclovir and alpha interferon (IFN-␣) prophylaxis, no KSHV-associated chemotherapy targets the underlying infection, a major oncogenic force. Hence, drugs that selectively target KSHV infection are necessary to eradicate the malignancy while sparing healthy cells. We recently showed that KSHV infection of endothelial cells activates the transcription factor Nrf2 to promote an environment conducive to infection and oncogenesis. Nrf2 is modulated through several well-tolerated oral agents and may be an important target in KSHV biology. Here, we investigate the role of Nrf2 in PEL and demonstrate that Nrf2 plays an important role in KSHV gene expression, lytic reactivation, and cell survival by interacting with the host transcriptional repressor KAP1 and the viral latency-...

show abstract

KSHV viral cyclin inactivates p27KIP1 through Ser10 and Thr187 phosphorylation in proliferating primary effusion lymphomas

Cited by 54 publications

References 49 publications

Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions

Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Multifunctional Angiogenin To Utilize Its Antiapoptotic Functions

Reactivation of the p53 pathway as a treatment modality for KSHV-induced lymphomas

Activated Nrf2 Interacts with Kaposi's Sarcoma-Associated Herpesvirus Latency Protein LANA-1 and Host Protein KAP1 To Mediate Global Lytic Gene Repression

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