Immunogenicity of and apoptosis modulation by Epstein-Barr virus (EBV)-encoded latent membrane protein-1 (LMP1) : implications for nasopharyngeal carcinoma

Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus which infects more than 90% of individuals in the population, and has been shown to be associated with a variety of human tumors. The viral genome encodes a number of transforming proteins which enable the virus to efficiently immortalize B lymphocyte and also transform cells of other types. Among the viral genetic products, a membrane integral latent infection protein, latent membrane protein-1 (LMP1) is of particular interest, since it possesses in vitro transforming ability and is expressed in more than two-thirds of EBV positive nasopharyngeal carcinoma (NPC), which arises in immunocompetent hosts.

Because of the distinctive geographic distribution and ethnic prevalence of NPC, strain variations of EBV carrying mutations in LMP1 have been investigated to correlate its presence in malignancy. Enhanced ability in transducing cellular signals as well as altered immunogenicity have been identified in LMP1 variant genes isolated from NPC specimens originating from the endemic regions. In the present study, a comparison of immunogenicity as related to the status of LMP1 expression in NPC biopsies has been made.

Our data (Paper I) show that LMP1 strains from LMP1-positive NPC contain more sequence variations than those from LMP1-negative NPC, and are less immunogenic. These results suggest a viral escape from host immune surveillance through genetic mutation in NPC. Modulation of host cell apoptosis is a strategy utilized by viruses to escape host immunity. LMP1 has been observed to reduce apoptosis in B and T cells, but its effect on apoptosis in epithelial cells remains obscure. We provide evidence herein (Paper II) for a stimulus-dependent apoptosis modulation by LMP1 in HeLa cells with tetracycline-regulated LMP1 expression; hence, LMP1 reduces tumor necrosis factor (TNF)-â induced apoptosis, but potentiates apoptosis induced upon ligation of Fas/Apo1/CD95, or following treatment with the chemotherapeutic agent, etoposide. It has recently reported that caspase-2 is an initiator caspase which regulates mitochondrial permeabilization, leading to the release of apoptosis inducing molecules into the cytoplasm.

In our model of stress-induced apoptosis, LMP1 promotes host cell apoptosis as manifested by enhancing the caspase-3 activity, and we have further shown that LMP1 regulates this type of apoptosis upstream of caspase-2 dependent mitochondrial perturbation. This effect was mapped to the membrane distal functional domain of LMP1, carboxylterminal activation region 2 (CTAR) (Paper III). The domain contributes to 70% of NF-kappaB activation and activates c-Jun N-terminal kinase (JNK) through recruitment of tumor necrosis factor receptor (TNFR) associated death domain containing molecule (TRADD).

We have shown that inhibition of either NF-kappaB or JNK activity regulates caspases and drop of mitochondrial membrane potential (MMP) triggered by cellular stress in LMP1 expressing HeLa cells (Paper IV). Furthermore LMP1 promotes irradiation-induced cell death assayed by staining of exposure of phosphatidylserine (PS) with annexin V in NPC derived TW03 cells, Future efforts will be aimed at further exploring the mechanism of LMP1-dependent modulation of cellular stress-mediated apoptosis, as well as the role thereof in the therapeutic response of EBV-associated epithelial human tumors.

List of scientific papers

I. Hu L, Troyanovsky B, Zhang X, Trivedi P, Ernberg I, Klein G (2000). Differences in the immunogenicity of latent membrane protein 1 (LMP1) encoded by Epstein-Barr virus genomes derived from LMP1-positive and -negative nasopharyngeal carcinoma. Cancer Res. 60(19): 5589-93.
https://pubmed.ncbi.nlm.nih.gov/11034108

II. Zhang X, Hu L, Fadeel B, Ernberg IT (2002). Apoptosis modulation of Epstein-Barr virus-encoded latent membrane protein 1 in the epithelial cell line HeLa is stimulus-dependent. Virology. 304(2): 330-41.
https://pubmed.ncbi.nlm.nih.gov/12504573

III. Zhang X, Uthaisang W, Hu L, Ernberg IT, Fadeel B (2005). Epstein-Barr virus-encoded latent membrane protein 1 promotes stress-induced apoptosis upstream of caspase-2-dependent mitochondrial perturbation. Int J Cancer. 113(3): 397-405.
https://pubmed.ncbi.nlm.nih.gov/15455353

IV. Zhang X, Sanmun D, Hu L, Fadeel B, Ernberg I (2005). Epstein-Barr virus-encoded latent membrane protein 1 (LMP1) promotes cisplatin-induced apoptosis through nuclear factor-kappaB and c-Jun-N-terminal kinase signaling pathways. [Manuscript]