[Construction and function analysis of the Epstein-Barr virus-encoded latent membrane protein-1 of CTAR3 region].
OBJECTIVE
The role of carboxyl terminal activating region of latent membrane protein1 (LMP1) in Epstein-Barr virus infection and oncogenesis is unclear. In this study, we investigated the activating sites and functionary mechanism of LMP1.


METHODS
We recombined a deletion mutant type LMP1 (LMP1Delta232-351), deleted the amino acid residues including 232-351 codons in carboxyl terminal activating region-3 by PCR. Then we compared mutant type LMP1Delta232-351 with wild type LMP1 (LMP1WT) to alter biological effect in Nasopharyngeal Epithelial Cell line NP69. Moreover, we constructed a Janus Kinase 3 (JKA3) promoter luciferase reporter system (pGL-2/JAK3-LUC). We respectively cotransfected the LMP1Delta232-351 and LMP1WT with promoter including NF-kappaB binding sequence or JAK3 promoter luciferase reporter into 293 cells (controlled with pLNSX vector), and compared their actions to activating promoters by results of luciferase activity assay.


RESULTS
(1) The colony forming number (CFN) of NP69-LMP1Delta232-351 cells significantly decreased to compare with CFN of NP69-LMP1WT (n=3, p<0.01). (2) LMP1WT was able to up-regulate the transcription activity of JAK3 promoter and the level of up-regulation was correlated with its concentration in Human embryonic kidney 293 cell line; while LMP1Delta232-351 was almost defective ability to activate the promoter.


CONCLUSIONS
The carboxyl terminal activating region-3 may be one of the most important function sites of LMP1, which involved in activating the JAK3 promoter and regulating the expression of JAK3 protein.
Zhang
Zhiwei
Z
Cancer Research Institute of Medical College, Nanhua University, Hengyang 421001, China. zhangzhiweichina@yahoo.com.cn
Zhang
Qiong
Q
Yu
Yanhui
Y
Ouyang
Yongmei
Y
He
Zhimin
Z
D002460:Cell Line; D020031:Epstein-Barr Virus Infections / Q000821:virology; D017930:Genes, Reporter; D004854:Herpesvirus 4, Human / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D053616:Janus Kinase 3 / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017384:Sequence Deletion; D015533:Transcriptional Activation; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
C037662:EBV-associated membrane antigen, Epstein-Barr virus; D014763:Viral Matrix Proteins; D053616:Janus Kinase 3