Stable expression of EBERs in immortalized nasopharyngeal epithelial cells confers resistance to apoptotic stress.
Epstein-Barr virus (EBV) infection is closely associated with the development of nasopharyngeal carcinoma (NPC). The EBV-encoded RNAs (EBERs) are the most abundant EBV transcripts (about 10(7) copies per cell) in EBV infected cells. However, the cellular function of EBER expression, particularly in nasopharyngeal epithelial cells, remains poorly understood. EBERs acquire secondary structures analogous to double-stranded RNA (dsRNA) and may bind to the double-stranded RNA-dependent protein kinase (PKR) and interfere with its function. Activation of PKR involves autophosphorylation resulting in protein synthesis inhibition and cellular apoptosis. Induction of cellular apoptosis by activation of PKR may be an antiviral response adopted by virally infected cells. We have examined the functional properties of EBER expression in an immortalized nasopharyngeal epithelial cell line (NP69). Expression of EBERs was achieved by transfecting the NP69 cells with an EBER-expressing plasmid, pESK10. The EBER-expressing NP69 cells attained a higher growth rate compared to cells transfected with control plasmid (pcDNA3). However, the EBER-expressing NP69 cells did not form colonies in soft agar and were non-tumorigenic in nude mice. To investigate if EBERs may protect the nasopharyngeal epithelial cells from apoptotic insults, we treated the EBER-expressing NP69 cells with a dsRNA analogue, poly(I).poly(C) (pIC), to activate PKR in cells and examined for their responses. Lower level of PKR phosphorylation and elevation of Bcl-2 were observed in EBER-expressing NP69 cells. In addition, other apoptotic markers including the cleaved forms of caspase-3 and poly(ADP)ribose polymerase (PARP) were found to be lower in EBER-expressing NP69 cells after treatment with pIC. Lower phosphorylation levels of p38 MAPK (mitogen-activated protein kinase) and c-jun were also observed in EBER-expressing NP cells. Our results suggest that EBER expression may confer an apoptotic-resistant phenotype in immortalized nasopharyngeal epithelial cells.
Wong
Hing Lok
HL
Cancer Biology Laboratory, Department of Anatomy, Faculty of Medicine, University of Hong Kong, 21 Sassoon Road, Pokfulam, Hong Kong SAR, China.
Wang
Xianghong
X
Chang
Raymond Chuen-Chung
RC
Jin
Dong-Yan
DY
Feng
Huichen
H
Wang
Qi
Q
Lo
Kwok-Wai
KW
Huang
Dolly P
DP
Yuen
Po Wing
PW
Takada
Kenzo
K
Wong
Yong Chuan
YC
Tsao
Sai Wah
SW
D000818:Animals; D017209:Apoptosis; D002461:Cell Line, Transformed; D017690:Cell Transplantation; D004847:Epithelial Cells / Q000821:virology; D020031:Epstein-Barr Virus Infections / Q000150:complications; D004854:Herpesvirus 4, Human / Q000302:isolation & purification*; D006801:Humans; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009305:Nasopharynx / Q000821:virology*; D010766:Phosphorylation; D012367:RNA, Viral / Q000032:analysis*; D014162:Transfection
D012367:RNA, Viral