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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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Induction of PUMA expression in C666-1 cells leads to apoptosis. (A) Western blot analysis of PARP. C666-1 cells were treatment with either 1.5 μg/ml adriamycin or 80 μM etoposide for 48 h. Expression of p53, PUMA-β, PARP, and β-actin was examined by Western blotting. (B) TUNEL analysis of apoptotic cells. TUNEL assays were performed on adriamycin- or etoposide-treated C666-1 cells by using confocal microscopy. Representative images are shown in Fig. S4 (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 = 0.035 (by Student's t test); #, P = 0.011.
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fig6: Induction of PUMA expression in C666-1 cells leads to apoptosis. (A) Western blot analysis of PARP. C666-1 cells were treatment with either 1.5 μg/ml adriamycin or 80 μM etoposide for 48 h. Expression of p53, PUMA-β, PARP, and β-actin was examined by Western blotting. (B) TUNEL analysis of apoptotic cells. TUNEL assays were performed on adriamycin- or etoposide-treated C666-1 cells by using confocal microscopy. Representative images are shown in Fig. S4 (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 = 0.035 (by Student's t test); #, P = 0.011.

Mentions: Interestingly, we found that the resistance to apoptosis occurred only in C666-1 cells treated with low-dose adriamycin (<1.2 μg/ml). Apoptosis was induced when the concentration of adriamycin was increased to 1.5 μg/ml or higher. Likewise, C666-1 cells treated with 80 μM etoposide also underwent apoptosis (Fig. 6). The manifestation of apoptosis in treated C666-1 cells was indicated by PARP cleavage (Fig. 6 A) and by terminal deoxynucleotidyl transferase–mediated dUTP-biotin nick end labeling (TUNEL) assay, which measures DNA fragmentation (Fig. 6 B and Fig. S4, available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). To investigate the underlying mechanism of apoptosis, we determined the expression profile of p53 and PUMA-β. Interestingly, both p53 and PUMA-β were induced significantly by high-dose adriamycin and etoposide (Fig. 6 A). Collectively, our results are compatible with the notion that the steady-state amount of PUMA dictates cellular sensitivity to proapoptotic agents. According to this model, when PUMA expression in C666-1 cells was effectively blocked by miR-BART5, apoptosis was prevented (Fig. 5). However, if the inhibitory effect of miR-BART5 was overcome by high-dose proapoptotic agents, apoptosis was triggered (Fig. 6).


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)

Induction of PUMA expression in C666-1 cells leads to apoptosis. (A) Western blot analysis of PARP. C666-1 cells were treatment with either 1.5 μg/ml adriamycin or 80 μM etoposide for 48 h. Expression of p53, PUMA-β, PARP, and β-actin was examined by Western blotting. (B) TUNEL analysis of apoptotic cells. TUNEL assays were performed on adriamycin- or etoposide-treated C666-1 cells by using confocal microscopy. Representative images are shown in Fig. S4 (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 = 0.035 (by Student's t test); #, P = 0.011.
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Related In: Results  -  Collection

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fig6: Induction of PUMA expression in C666-1 cells leads to apoptosis. (A) Western blot analysis of PARP. C666-1 cells were treatment with either 1.5 μg/ml adriamycin or 80 μM etoposide for 48 h. Expression of p53, PUMA-β, PARP, and β-actin was examined by Western blotting. (B) TUNEL analysis of apoptotic cells. TUNEL assays were performed on adriamycin- or etoposide-treated C666-1 cells by using confocal microscopy. Representative images are shown in Fig. S4 (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 = 0.035 (by Student's t test); #, P = 0.011.
Mentions: Interestingly, we found that the resistance to apoptosis occurred only in C666-1 cells treated with low-dose adriamycin (<1.2 μg/ml). Apoptosis was induced when the concentration of adriamycin was increased to 1.5 μg/ml or higher. Likewise, C666-1 cells treated with 80 μM etoposide also underwent apoptosis (Fig. 6). The manifestation of apoptosis in treated C666-1 cells was indicated by PARP cleavage (Fig. 6 A) and by terminal deoxynucleotidyl transferase–mediated dUTP-biotin nick end labeling (TUNEL) assay, which measures DNA fragmentation (Fig. 6 B and Fig. S4, available at http://www.jem.org/cgi/content/full/jem.20072581/DC1). To investigate the underlying mechanism of apoptosis, we determined the expression profile of p53 and PUMA-β. Interestingly, both p53 and PUMA-β were induced significantly by high-dose adriamycin and etoposide (Fig. 6 A). Collectively, our results are compatible with the notion that the steady-state amount of PUMA dictates cellular sensitivity to proapoptotic agents. According to this model, when PUMA expression in C666-1 cells was effectively blocked by miR-BART5, apoptosis was prevented (Fig. 5). However, if the inhibitory effect of miR-BART5 was overcome by high-dose proapoptotic agents, apoptosis was triggered (Fig. 6).

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