Multiple sclerosis (MS) is a widespread neurodegenerative autoimmune disease with unknown etiology. It is increasingly evident that, together with pathogenic T cells, autoreactive B cells are among the major players in MS development. The analysis of myelin neuroantigen-specific antibody repertoires and their possible cross-reactivity against environmental antigens, including viral proteins, could shed light on the mechanism of MS induction and progression. A phage display library of single-chain variable fragments (scFvs) was constructed from blood lymphocytes of patients with MS as a potential source of representative MS autoantibodies. Structural alignment of 13 clones selected toward myelin basic protein (MBP), one of the major myelin antigens, showed high homology within variable regions with cerebrospinal fluid MS-associated antibodies as well as with antibodies toward Epstein-Barr latent membrane protein 1 (LMP1). Three scFv clones showed pronounced specificity to MBP fragments 65-92 and 130-156, similar to the serum MS antibodies. One of these clones, designated E2, in both scFv and full-size human antibody constructs, was shown to react with both MBP and LMP1 proteins in vitro, suggesting natural cross-reactivity. Thus, antibodies induced against LMP1 during Epstein-Barr virus infection might act as inflammatory trigger by reacting with MBP, suggesting molecular mimicry in the mechanism of MS pathogenesis.

The genetic structure of EBV LMP1 alleles isolated from tumor, blood, and throat washing samples of 22 nasopharyngeal carcinoma patients, 17 patients with other non-EBV-related tumors of the oral cavity, and 19 blood donors have been studied in representatives of Central Russia and the Republics of Northern Caucasus, regions which are non-endemic for nasopharyngeal carcinoma. The analysis of the LMP1 alleles collected revealed that they practically matched previously described LMP1 variants; however, some characteristic features were also detected. In particular, the G212S substitution in LMP1 isolates investigated was not observed at all. Tumor samples obtained from nasopharyngeal carcinoma and other tumors of the oral cavity did not differ significantly either in the frequency of "high oncogenic" LMP1 alleles with 10 aa and/or 23 aa deletions (LMP1(China1) and/or LMP1(Med+)), nor in the number of 11 aa repeats and the frequency of 5 aa motif insertions. No differences in the frequency of amino acid substitutions between LMP1 alleles obtained from tumor and throat washing samples of both patient groups were also detected. The data obtained may indicate that in both nasopharyngeal carcinoma patients and patients with other tumors of the oral cavity, the EBV strains with similar LMP1 variants are found to persist. This observation allows us to suggest that in non-endemic areas, EBV strains with any LMP1 alleles can initiate the nasopharyngeal carcinoma development but only in those individuals who have a genetic predisposition to the disease and are subjected to specific environmental, and/or dietary factors present in certain geographic areas.

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Latent membrane protein 1 (LMP1) of the Epstein-Barr virus is a constitutively activated analog of the tumor necrosis factor receptor TNF-R1. LMP1 serves as a viral oncogene able to transform human B-lymphocytes and rodent fibroblasts via activation of numerous cellular signal cascades. Two specific motifs within LMP1 are responsible for interaction of this viral protein with the receptor protein beta-TrCP/HOS SCF of the ubiquitin ligase E3 complex, playing an important role in degradation of numerous cellular proteins including NF-kappaB inhibitor IkappaBalpha. In this study, we demonstrate for the first time the importance of point mutations affecting HOS-recognizing motifs of LMP1 for activation of NF-kappaB, AP1, and PI3K/Akt signaling pathways. It has also been shown that rat fibroblast cell lines (Rat-1) expressing different HOS mutants of LMP1 produce different amounts of reactive nitrogen species. Our data confirm the hypothesis that point mutations in the C-terminal region of the LMP1 cytoplasmic domain can influence the transforming potential of the Epstein-Barr virus.

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Latent membrane protein1 (LMP1) encoded by the LMP1 gene of the Epstein-Barr virus (EBV) is a transmembrane protein, which can activate a number of cellular signal cascades and transcriptional factors leading to cell transformation. In the present study the sequencing of full-length LMP1 variants isolated from Russian patients with Hodgkin's lymphoma (HL), non-Hodgkin's lymphomaе (NHL) and infectious mononucleosis (IM) has been carried out. The phylogenetic analysis of obtained sequences revealed dominance of the LMP1 variants belonging to proteins of low-divergent group LMP1-B95.8b characterized by minimal set of mutations. Investigation of biological properties in the Russian representatives of this group revealed that expression of studied LMP1 variants in embryonic kidney 293 cells was accompanied by insignificant increase in transcriptional factor NF-κB activation and had minor influence on activation of transcriptional factor AP-1. It was also detected that all investigated low-divergent LMP1 variants expressed in Rat-1 cells induced activation of inducible NO-synthase (iNOS) and intracellular production of nitric oxide (NO). At the same time the level of NO accumulation was lower than that induced by the low-transforming prototype variant LMP1-B95.8. The data obtained indicate that the LMP1 variants, which are the most common among Russian patients with EBV-associated lymphoproliferative diseases, are characterized by minimum number of mutations and rather low ability to activate basic cellular signaling pathways regardless the nature of pathological process, benign (IM) or malignant (HL, NHL). It is suggested that in addition to the modest activation of NF-κB and iNOS induction by LMP1 other factors are involved in the cell transformation process.

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