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An Epstein-Barr virus-encoded microRNA targets PUMA to promote host cell survival.

Choy EY, Siu KL, Kok KH, Lung RW, Tsang CM, To KF, Kwong DL, Tsao SW, Jin DY - J. Exp. Med. (2008)

Bottom Line: In addition, PUMA was found to be significantly underexpressed in approximately 60% of human NPC tissues.Although expression of miR-BART5 rendered NPC and EBV-GC cells less sensitive to proapoptotic agents, apoptosis can be triggered by depleting miR-BART5 or inducing the expression of PUMA.Collectively, our findings suggest that EBV encodes an miRNA to facilitate the establishment of latent infection by promoting host cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, The University of Hong Kong, Pokfulam, Hong Kong.

ABSTRACT
Epstein-Barr virus (EBV) is a herpesvirus associated with nasopharyngeal carcinoma (NPC), gastric carcinoma (GC), and other malignancies. EBV is the first human virus found to express microRNAs (miRNAs), the functions of which remain largely unknown. We report on the regulation of a cellular protein named p53 up-regulated modulator of apoptosis (PUMA) by an EBV miRNA known as miR-BART5, which is abundantly expressed in NPC and EBV-GC cells. Modulation of PUMA expression by miR-BART5 and anti-miR-BART5 oligonucleotide was demonstrated in EBV-positive cells. In addition, PUMA was found to be significantly underexpressed in approximately 60% of human NPC tissues. Although expression of miR-BART5 rendered NPC and EBV-GC cells less sensitive to proapoptotic agents, apoptosis can be triggered by depleting miR-BART5 or inducing the expression of PUMA. Collectively, our findings suggest that EBV encodes an miRNA to facilitate the establishment of latent infection by promoting host cell survival.

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Inhibition of miR-BART5 in C666-1 cells induces apoptosis. (A) Induction of apoptosis. C666-1 cells were transfected with anti–miR-BART5 or anti–miRNA-NC oligonucleotide inhibitor. Cells were treated with etoposide as in Fig. 6 and TUNEL assay was performed. Representative images are shown in Fig. S5 (available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). 250 cells were scored and the quantitative results represent mean ± SD from three independent experiments. *, P = 0.03 (by Student's t test); #, P = 0.05. (B) Suppression of PUMA expression by siRNA. C666-1 cells were transfected with 100 nM siGFP or siPUMA. Expression of PUMA-β was examined by Western blotting. (C) Inhibition of miR-BART5 activity by siPUMA. C666-1 cells were cotransfected with anti–miR-BART5 and siPUMA. Apoptotic cells were analyzed as in A. Error bars indicate SD (n = 3).
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fig7: Inhibition of miR-BART5 in C666-1 cells induces apoptosis. (A) Induction of apoptosis. C666-1 cells were transfected with anti–miR-BART5 or anti–miRNA-NC oligonucleotide inhibitor. Cells were treated with etoposide as in Fig. 6 and TUNEL assay was performed. Representative images are shown in Fig. S5 (available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). 250 cells were scored and the quantitative results represent mean ± SD from three independent experiments. *, P = 0.03 (by Student's t test); #, P = 0.05. (B) Suppression of PUMA expression by siRNA. C666-1 cells were transfected with 100 nM siGFP or siPUMA. Expression of PUMA-β was examined by Western blotting. (C) Inhibition of miR-BART5 activity by siPUMA. C666-1 cells were cotransfected with anti–miR-BART5 and siPUMA. Apoptotic cells were analyzed as in A. Error bars indicate SD (n = 3).

Mentions: To further strengthen this model, we asked whether inhibition of miR-BART5 by another means would also induce apoptosis. Because miR-BART5 can be specifically and effectively inhibited by anti–miR-BART5 oligonucleotide (Fig. 4, C and D), we tested the influence of this miR-BART5 inhibitor on apoptosis of C666-1 and AGS/BX1 cells. Notably, transfection of C666-1 cells with anti–miR-BART5 oligonucleotide triggered apoptosis (Fig. 7 A and Fig. S5, available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). Additionally, anti–miR-BART5 oligonucleotide was able to enhance the proapoptotic effect of etoposide mildly leading to an ∼10% increase of TUNEL-positive apoptotic cells (Fig. 7 A). Plausibly, both anti–miR-BART5 oligonucleotide and etoposide exert their proapoptotic effects by inducing the expression of PUMA.


An Epstein-Barr virus-encoded microRNA targets PUMA to promote host cell survival.

Choy EY, Siu KL, Kok KH, Lung RW, Tsang CM, To KF, Kwong DL, Tsao SW, Jin DY - J. Exp. Med. (2008)

Inhibition of miR-BART5 in C666-1 cells induces apoptosis. (A) Induction of apoptosis. C666-1 cells were transfected with anti–miR-BART5 or anti–miRNA-NC oligonucleotide inhibitor. Cells were treated with etoposide as in Fig. 6 and TUNEL assay was performed. Representative images are shown in Fig. S5 (available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). 250 cells were scored and the quantitative results represent mean ± SD from three independent experiments. *, P = 0.03 (by Student's t test); #, P = 0.05. (B) Suppression of PUMA expression by siRNA. C666-1 cells were transfected with 100 nM siGFP or siPUMA. Expression of PUMA-β was examined by Western blotting. (C) Inhibition of miR-BART5 activity by siPUMA. C666-1 cells were cotransfected with anti–miR-BART5 and siPUMA. Apoptotic cells were analyzed as in A. Error bars indicate SD (n = 3).
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fig7: Inhibition of miR-BART5 in C666-1 cells induces apoptosis. (A) Induction of apoptosis. C666-1 cells were transfected with anti–miR-BART5 or anti–miRNA-NC oligonucleotide inhibitor. Cells were treated with etoposide as in Fig. 6 and TUNEL assay was performed. Representative images are shown in Fig. S5 (available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). 250 cells were scored and the quantitative results represent mean ± SD from three independent experiments. *, P = 0.03 (by Student's t test); #, P = 0.05. (B) Suppression of PUMA expression by siRNA. C666-1 cells were transfected with 100 nM siGFP or siPUMA. Expression of PUMA-β was examined by Western blotting. (C) Inhibition of miR-BART5 activity by siPUMA. C666-1 cells were cotransfected with anti–miR-BART5 and siPUMA. Apoptotic cells were analyzed as in A. Error bars indicate SD (n = 3).
Mentions: To further strengthen this model, we asked whether inhibition of miR-BART5 by another means would also induce apoptosis. Because miR-BART5 can be specifically and effectively inhibited by anti–miR-BART5 oligonucleotide (Fig. 4, C and D), we tested the influence of this miR-BART5 inhibitor on apoptosis of C666-1 and AGS/BX1 cells. Notably, transfection of C666-1 cells with anti–miR-BART5 oligonucleotide triggered apoptosis (Fig. 7 A and Fig. S5, available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). Additionally, anti–miR-BART5 oligonucleotide was able to enhance the proapoptotic effect of etoposide mildly leading to an ∼10% increase of TUNEL-positive apoptotic cells (Fig. 7 A). Plausibly, both anti–miR-BART5 oligonucleotide and etoposide exert their proapoptotic effects by inducing the expression of PUMA.

Bottom Line: In addition, PUMA was found to be significantly underexpressed in approximately 60% of human NPC tissues.Although expression of miR-BART5 rendered NPC and EBV-GC cells less sensitive to proapoptotic agents, apoptosis can be triggered by depleting miR-BART5 or inducing the expression of PUMA.Collectively, our findings suggest that EBV encodes an miRNA to facilitate the establishment of latent infection by promoting host cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, The University of Hong Kong, Pokfulam, Hong Kong.

ABSTRACT
Epstein-Barr virus (EBV) is a herpesvirus associated with nasopharyngeal carcinoma (NPC), gastric carcinoma (GC), and other malignancies. EBV is the first human virus found to express microRNAs (miRNAs), the functions of which remain largely unknown. We report on the regulation of a cellular protein named p53 up-regulated modulator of apoptosis (PUMA) by an EBV miRNA known as miR-BART5, which is abundantly expressed in NPC and EBV-GC cells. Modulation of PUMA expression by miR-BART5 and anti-miR-BART5 oligonucleotide was demonstrated in EBV-positive cells. In addition, PUMA was found to be significantly underexpressed in approximately 60% of human NPC tissues. Although expression of miR-BART5 rendered NPC and EBV-GC cells less sensitive to proapoptotic agents, apoptosis can be triggered by depleting miR-BART5 or inducing the expression of PUMA. Collectively, our findings suggest that EBV encodes an miRNA to facilitate the establishment of latent infection by promoting host cell survival.

Show MeSH
Related in: MedlinePlus