Protein complexes associated with the Kaposi's sarcoma-associated herpesvirus-encoded LANA

Kaul, Rajeev; Verma, Subhash C.; Robertson, Erle S.

doi:10.1016/j.virol.2007.03.010

Cited by 48 publications

(69 citation statements)

References 72 publications

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“…For expression of GST and GST fusion proteins, bacterial culture was induced with 1 mM isopropyl-␤-D-thiogalactopyranoside (IPTG) at log phase (optical density at 600 nm [OD 600 ] ϭ 0.6), and cells were incubated with shaking at 37°C for 4 h. Bacterial lysis and purification of protein(s) with glutathione-Sepharose beads were carried out as described earlier (29). Quantification was done by SDS-PAGE of purified proteins with bovine serum albumin (BSA) standards followed by densitometry analysis of Coomassie brilliant blue-stained gel using an Odyssey imager (LiCor Inc., Lincoln, NE).…”

Section: Methodsmentioning

confidence: 99%

H2AX Phosphorylation Is Important for LANA-Mediated Kaposi's Sarcoma-Associated Herpesvirus Episome Persistence

Jha

Upadhyay

et al. 2013

J Virol

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The DNA damage response (DDR) of host cells is utilized by a number of viruses to establish and propagate their genomes in the infected cells. We examined the expression of the DDR genes during Kaposi's sarcoma-associated herpesvirus (KSHV) infection of human peripheral blood mononuclear cells (PBMCs). The genes were mostly downregulated, except H2AX, which was upregulated during infection. H2AX is important for gammaherpesvirus infectivity, and its phosphorylation at serine 139 is crucial for maintenance of latency during mouse gamma-herpesvirus 68 (MHV-68) infection. We now also observed phosphorylation of H2AX at serine 139 during KSHV infection. H2AX is a histone H2A isoform shown to interact with the latency-associated nuclear antigen (LANA) encoded by KSHV. Here, we show that LANA directly interacted with H2AX through domains at both its N and C termini. The phosphorylated form of H2AX (␥H2AX) was shown to colocalize with LANA. Chromatin immunoprecipitation (ChIP) assays showed that a reduction in H2AX levels resulted in reduced binding of LANA with KSHV terminal repeats (TRs). Binding preferences of H2AX and ␥H2AX along the KSHV episome were examined by whole-episome ChIP analysis. We showed that ␥H2AX had a higher relative binding activity along the TR regions than that of the long unique region (LUR), which highlighted the importance

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Section: Methodsmentioning

confidence: 99%

H2AX Phosphorylation Is Important for LANA-Mediated Kaposi's Sarcoma-Associated Herpesvirus Episome Persistence

Jha

Upadhyay

et al. 2013

J Virol

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“…A yeast two- hybrid screen, using the carboxy terminus of LANA as bait with a cDNA library from lymphoblastoid cell lines, identified NuMA as a candidate LANA-interacting molecule. Additionally, recent data from a proteomic study using the carboxy terminus of LANA also detected NuMA, suggesting strong association of NuMA with LANA (31). To further corroborate the interaction, we performed GST-binding assays using vector, amino terminus, and the carboxy terminus of LANA fused to GST (Fig.…”

Section: Resultsmentioning

confidence: 69%

“…This is the first report showing that a viral protein can interact with NuMA and that it helps in segregation of the viral genome into the dividing daughter cells. A role for KSHV LANA protein in attaching the viral genome to the nuclear matrix was shown before by our group, in that CENP-F binds to LANA and helps in tethering the viral genome to the nuclear matrix (31). A human papillomavirus 16-encoded protein, E7, has also been shown to colocalize with the nuclear matrix during infection (23).…”

Section: Vol 82 2008mentioning

confidence: 63%

“…Dyneins are involved in the transport of particles and organelles along microtubules and in the transport of condensed chromosomes during mitosis (18). In our previous proteomic studies, heavy-chain dyneins have been found to bind both the KSHV TR and LANA (31,56). Since dynein/dynactin and microtubule association are important in transportation of both NuMA and mitotic chromosomes to the spindle poles, we speculate that NuMA may also help in efficient segregation of the replicated KSHV genome into daughter cells.…”

Section: Vol 82 2008mentioning

confidence: 74%

“…In somatic cells, this interaction is restricted to mitosis, because NuMA is segregated to the nucleus during interphase and spatially separated from cytoplasmic dynein and dynactin by the nuclear membrane. Additionally, previous proteomic studies from our laboratory have shown that dynein can bind to LANA (31,56). Thus, we wanted to see if these proteins were functionally involved in KSHV genome persistence.…”

Section: Vol 82 2008mentioning

confidence: 99%

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Kaposi's Sarcoma-Associated Herpesvirus-Encoded LANA Can Interact with the Nuclear Mitotic Apparatus Protein To Regulate Genome Maintenance and Segregation

Verma

Lampson

et al. 2008

J Virol

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Kaposi's sarcoma-associated herpesvirus (KSHV) genomes are tethered to the host chromosomes and partitioned faithfully into daughter cells with the host chromosomes. The latency-associated nuclear antigen (LANA) is important for segregation of the newly synthesized viral genomes to the daughter nuclei. Here, we report that the nuclear mitotic apparatus protein (NuMA) and LANA can associate in KSHV-infected cells. In synchronized cells, NuMA and LANA are colocalized in interphase cells and separate during mitosis at the beginning of prophase, reassociating again at the end of telophase and cytokinesis. Silencing of NuMA expression by small interfering RNA and expression of LGN and a dominant-negative of dynactin (P150-CC1), which disrupts the association of NuMA with microtubules, resulted in the loss of KSHV terminal-repeat plasmids containing the major latent origin. Thus, NuMA is required for persistence of the KSHV episomes in daughter cells. This interaction between NuMA and LANA is critical for segregation and maintenance of the KSHV episomes through a temporally controlled mechanism of binding and release during specific phases of mitosis.

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Proteomic profiling identifies the SIM‐associated complex of KSHV‐encoded LANA

et al. 2015

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The Kaposi’s sarcoma-associated herpesvirus (KSHV)-encoded latent nuclear antigen LANA plays an essential role in viral episome maintenance. LANA also contributes to DNA replication and tumorigenesis during latency. Recent studies suggested that LANA was involved in regulation of SUMOylation which results in chromatin silencing. To examine the pleiotropic effects of LANA protein on host cell gene expression, we utilized MS analysis to identify cellular proteins associated with the SUMO-Interacting Motif of LANA (LANASIM). In addition to the 6 bands identified as substantially associated with LANASIM, 151 proteins were positively identified by MS/MS analysis. Compared with previous proteomic analysis of the N- and C- truncated mutants of LANA (LANANC), our results revealed that a complex of specific proteins with relatively high SUMOylation and SIM motifs are associated with LANASIM. Intriguingly, consistent with our previous report that identified KAP1 as a key component, the in-vitro SUMO-2 modified isoform has a substantially higher affinity with LANASIM than the SUMO-1 modified isoform. Moreover, via cluster and pathway analysis, we proposed a hypothetical model for the LANASIM regulatory circuit involving aberrant SUMOylation of cell cycle (particular mitotic), DNA unwinding and replication, and pre-mRNA/mRNA processing–related proteins. This study provides a SUMOylated and non-SUMOylated proteome profile of LANASIM-associated complex, and facilitates our understanding that viral-mediated gene regulation through SUMOylation is important for KSHV persistence and pathogenesis.

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Protein complexes associated with the Kaposi's sarcoma-associated herpesvirus-encoded LANA

Cited by 48 publications

References 72 publications

H2AX Phosphorylation Is Important for LANA-Mediated Kaposi's Sarcoma-Associated Herpesvirus Episome Persistence

H2AX Phosphorylation Is Important for LANA-Mediated Kaposi's Sarcoma-Associated Herpesvirus Episome Persistence

Kaposi's Sarcoma-Associated Herpesvirus-Encoded LANA Can Interact with the Nuclear Mitotic Apparatus Protein To Regulate Genome Maintenance and Segregation

Proteomic profiling identifies the SIM‐associated complex of KSHV‐encoded LANA

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