Epstein-Barr virus (EBV) is a ubiquitous human herpesvirus that is associated with the development of lymphoid and epithelial malignancies including Hodgkin lymphoma (HD), post-transplant lymphoproliferative disorder (PTLD), nasopharyngeal carcinoma (NPC), and others. Latent membrane protein 1 (LMP1) is the EBV oncogene as it is necessary for EBV-mediated transformation of B-lymphocytes and can itself transform rodent fibroblasts. LMP1 has a short amino-terminal cytoplasmic end, 6 transmembrane domains, and a long carboxyl-terminal cytoplasmic tail that contains the two signaling domains, the carboxyl-terminal activation regions 1 and 2 (CTAR1 and CTAR2). Through CTAR1 and CTAR2, LMP1 activates several signaling pathways including NF[kappa]B, phosphatidylinositol 3-kinase (PI3K), mitogen-activated protein kinase (MAPK), and c-Jun N-terminal kinase (JNK) pathways. In this study, CTAR1 was shown to be necessary for Rat-1 fibroblast transformation and activation of the PI3K-Akt pathway, whereas CTAR2 was dispensable. LMP1-mediated rodent fibroblast transformation did not require I[kappa]B[alpha]-dependent activation of NF[kappa]B. Seven sequence variants of LMP1 have been identified: Alaskan, B95.8, China 1, China 2, Med+, Med-, and NC. The frequency of detection of these variants differs in various EBV-associated malignancies from different geographic regions. In this study, all the sequence variants were demonstrated to transform Rat-1 fibroblasts, induce increased motility of human foreskin keratinocytes, induce homotypic adhesion of BJAB cells and activate the PI3K-Akt signaling pathway. The Alaskan, China 1 and Med+ variants had limited binding to the E3 ubiquitin ligase component HOS and enhanced NF[kappa]B signaling. These findings indicate that the signature amino acid changes of the LMP1 variants do not hinder or significantly enhance their in vitro transforming potential or signaling properties. Finally, dominant negative forms of TRAF2 and TRAF3 inhibited LMP1-mediated transformation of rodent fibroblasts. Activation of PI3K-Akt signaling and deregulation of cell-cycle markers were mapped to the TRAF-binding site and residues between CTAR1 and CTAR2. Activation of the MEK1/2-ERK1/2 signaling pathway through the TRAF-binding site was required for LMP1-induced transformation of rodent fibroblasts. These findings identify a signaling pathway that is uniquely activated by CTAR1.