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TRIM52 inhibits Japanese Encephalitis Virus replication by degrading the viral NS2A

View Article: PubMed Central - PubMed

ABSTRACT

The members of tripartite-motif containing (TRIM) protein participate in various cellular processes and play an important role in host antiviral function. TRIM proteins exert their antiviral activity either directly by degrading viral proteins through their E3 ligase activity, or indirectly by promoting host innate immunity. This study demonstrated for the first time that TRIM52 is a novel antiviral TRIM protein against Japanese encephalitis virus (JEV) infection. Overexpression of TRIM52 restricted JEV replication in BHK-21 and 293T cells. In addition, JEV nonstructural protein 2A (NS2A) is a protein that interacts with TRIM52. Their interaction degraded NS2A in a proteasome-dependent manner via the E3 ligase activity of TRIM52. Thus, TRIM52 is a novel antiviral TRIM protein, and it exerted antiviral activity against JEV infection by targeting and degrading viral NS2A.

No MeSH data available.


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TRIM52 expression impairs JEV infection in BHK-21 and 293T cells.The BHK-21 or 293T cells in 12-well plates were transfected with 2 μg of TRIM52 expression plasmids or with empty vector for 30 h. The cells were subsequently infected with JEV at an MOI of 1. The cells and supernatants were collected at 24 h post-infection. Indicated experiments were performed to evaluate the antiviral activity of TRIM52 against JEV infection. (a) Production of progeny virus in culture supernatants of BHK-21 (left) and 293T cells (right) was measured by plaque assay. (b) Absolute quantification of intracellular JEV C gene copies in BHK-21 (left) and 293T cells (right) through real-time PCR analysis. (c) Western blot analysis of the protein level of JEV NS3 in BHK-21 (left) and 293T cells (right) by using the indicated antibodies. GAPDH served as loading control at an equal sample loading. Three asterisks indicate that a statistical difference exists between empty vector and TRIM52 expression cells at P < 0.001.
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f1: TRIM52 expression impairs JEV infection in BHK-21 and 293T cells.The BHK-21 or 293T cells in 12-well plates were transfected with 2 μg of TRIM52 expression plasmids or with empty vector for 30 h. The cells were subsequently infected with JEV at an MOI of 1. The cells and supernatants were collected at 24 h post-infection. Indicated experiments were performed to evaluate the antiviral activity of TRIM52 against JEV infection. (a) Production of progeny virus in culture supernatants of BHK-21 (left) and 293T cells (right) was measured by plaque assay. (b) Absolute quantification of intracellular JEV C gene copies in BHK-21 (left) and 293T cells (right) through real-time PCR analysis. (c) Western blot analysis of the protein level of JEV NS3 in BHK-21 (left) and 293T cells (right) by using the indicated antibodies. GAPDH served as loading control at an equal sample loading. Three asterisks indicate that a statistical difference exists between empty vector and TRIM52 expression cells at P < 0.001.

Mentions: Many members of the TRIM family act as regulator in host–virus interaction; they either positively or negatively impact virus replication, as well as directly or indirectly restrict virus replication. This study investigated the potential antiviral function of TRIM52 against JEV infection. To test whether TRIM52 can influence JEV replication, we transfected BHK-21 and 293T cells with plasmids expressing HA-tagged TRIM52 or with an empty vector as control. The control and TRIM52–overexpressing cells were both infected with JEV at an MOI of 1. The culture supernatants were harvested 24 hour post-infection (hpi), and the titers of the progeny virus were determined using plaque forming unit assay on BHK-21 cells. Figure 1a shows that TRIM52 overexpression reduced JEV titers both in BHK-21 (left panel) and 293T cells (right panel). Moreover, gene copies of JEV C gene and protein level of NS3 in infected cells were detected, demonstrating the antiviral activity of TRIM52 against JEV. As shown in Fig. 1b, the intracellular C gene levels of JEV were significantly reduced in TRIM52-overexpressing cells compared with that in the vector control cells. In addition, Western blot analysis indicated that the NS3 protein levels of JEV were significantly repressed in the presence of TRIM52. These results suggest that TRIM52 exerted antiviral activity against JEV replication both in BHK-21 and 293T cells.


TRIM52 inhibits Japanese Encephalitis Virus replication by degrading the viral NS2A
TRIM52 expression impairs JEV infection in BHK-21 and 293T cells.The BHK-21 or 293T cells in 12-well plates were transfected with 2 μg of TRIM52 expression plasmids or with empty vector for 30 h. The cells were subsequently infected with JEV at an MOI of 1. The cells and supernatants were collected at 24 h post-infection. Indicated experiments were performed to evaluate the antiviral activity of TRIM52 against JEV infection. (a) Production of progeny virus in culture supernatants of BHK-21 (left) and 293T cells (right) was measured by plaque assay. (b) Absolute quantification of intracellular JEV C gene copies in BHK-21 (left) and 293T cells (right) through real-time PCR analysis. (c) Western blot analysis of the protein level of JEV NS3 in BHK-21 (left) and 293T cells (right) by using the indicated antibodies. GAPDH served as loading control at an equal sample loading. Three asterisks indicate that a statistical difference exists between empty vector and TRIM52 expression cells at P < 0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5035999&req=5

f1: TRIM52 expression impairs JEV infection in BHK-21 and 293T cells.The BHK-21 or 293T cells in 12-well plates were transfected with 2 μg of TRIM52 expression plasmids or with empty vector for 30 h. The cells were subsequently infected with JEV at an MOI of 1. The cells and supernatants were collected at 24 h post-infection. Indicated experiments were performed to evaluate the antiviral activity of TRIM52 against JEV infection. (a) Production of progeny virus in culture supernatants of BHK-21 (left) and 293T cells (right) was measured by plaque assay. (b) Absolute quantification of intracellular JEV C gene copies in BHK-21 (left) and 293T cells (right) through real-time PCR analysis. (c) Western blot analysis of the protein level of JEV NS3 in BHK-21 (left) and 293T cells (right) by using the indicated antibodies. GAPDH served as loading control at an equal sample loading. Three asterisks indicate that a statistical difference exists between empty vector and TRIM52 expression cells at P < 0.001.
Mentions: Many members of the TRIM family act as regulator in host–virus interaction; they either positively or negatively impact virus replication, as well as directly or indirectly restrict virus replication. This study investigated the potential antiviral function of TRIM52 against JEV infection. To test whether TRIM52 can influence JEV replication, we transfected BHK-21 and 293T cells with plasmids expressing HA-tagged TRIM52 or with an empty vector as control. The control and TRIM52–overexpressing cells were both infected with JEV at an MOI of 1. The culture supernatants were harvested 24 hour post-infection (hpi), and the titers of the progeny virus were determined using plaque forming unit assay on BHK-21 cells. Figure 1a shows that TRIM52 overexpression reduced JEV titers both in BHK-21 (left panel) and 293T cells (right panel). Moreover, gene copies of JEV C gene and protein level of NS3 in infected cells were detected, demonstrating the antiviral activity of TRIM52 against JEV. As shown in Fig. 1b, the intracellular C gene levels of JEV were significantly reduced in TRIM52-overexpressing cells compared with that in the vector control cells. In addition, Western blot analysis indicated that the NS3 protein levels of JEV were significantly repressed in the presence of TRIM52. These results suggest that TRIM52 exerted antiviral activity against JEV replication both in BHK-21 and 293T cells.

View Article: PubMed Central - PubMed

ABSTRACT

The members of tripartite-motif containing (TRIM) protein participate in various cellular processes and play an important role in host antiviral function. TRIM proteins exert their antiviral activity either directly by degrading viral proteins through their E3 ligase activity, or indirectly by promoting host innate immunity. This study demonstrated for the first time that TRIM52 is a novel antiviral TRIM protein against Japanese encephalitis virus (JEV) infection. Overexpression of TRIM52 restricted JEV replication in BHK-21 and 293T cells. In addition, JEV nonstructural protein 2A (NS2A) is a protein that interacts with TRIM52. Their interaction degraded NS2A in a proteasome-dependent manner via the E3 ligase activity of TRIM52. Thus, TRIM52 is a novel antiviral TRIM protein, and it exerted antiviral activity against JEV infection by targeting and degrading viral NS2A.

No MeSH data available.


Related in: MedlinePlus