SEQUENCE VARIATIONS BETWEEN 2 EPSTEIN-BARR-VIRUS LMP-1 VARIANTS HAVE NO EFFECT ON THE ACTIVATION OF NF-KAPPA-B ACTIVITY

Previously, we reported that the LMP 1 gene of Epstein-Barr virus (EBV ) derived from nasopharyngeal carcinoma (NPC) tissues (i.e., NLMP 1 ge ne) was able to transform BALB/c3T3 cells, On the other hand, LMP I ge ne of B95-8 strain (i.e., BLMP 1 gene) was not able to transform these cells (Chen et al., 1992), Further studies indicated that a 10-amino- acid deletion in the carboxyl terminus of NLMP 1 played an important r ole in transformation (Li et al., 1996), In this study, we tested if t his 10-amino-acid deletion affected the induction of NF-kappa B activi ty by LMP 1, The long terminal repeat of the human immunodeficiency vi rus type 1 (HIV-1 LTR) contained two copies of NF-kappa B sites and wa s used to construct the Luc gene-based reporter plasmid, p kappa B-Luc , Plasmid p kappa B-Luc was co-transfected with plasmids containing th e NLMP 1 gene, BLMP 1 gene, and their chimeric or deletion constructs, respectively, into C-33A and BALB/c3T3 cells, The activation was then measured by the luciferase activity, Results showed that the full-len gth proteins induced a similar level of NF-kappa B activity, the two 3 ' mutants (R15 Delta and D4 Delta) still induced a relatively high lev el of activity, and the two 5' deletion mutants (Delta 3058 and Delta 3243) of NLMP 1 gene did not show any significant activation in C-33A cells, However, none of these LMP 1 proteins induced NF-kappa B activi ty in BALB/c3T3 cells, Using subcellular fractionation analysis and an immunocytostaining method, the truncated proteins of Delta 3058 and D elta 3243 were detected in the cytoplasm of the cells whereas the full -length NLMP 1 protein was located at the cytoplasmic membrane, Stable BALB/c3T3 cell clones that expressed both truncated proteins were est ablished and then their ability to induce tumors in nude mice was exam ined, Data showed that both truncated NLMP 1 proteins still maintained partial transformation activity, Our results suggested that there was no direct correlation between NF-kappa B activation and transformatio n activity of LMP 1 in BALB/c3T3 cell transformation and that the amin o-terminal membrane-spanning domain was important for maintaining both functions of LMP 1.