Novel Patterns of the Epstein-Barr Nuclear Antigen (EBNA-1) V-Val Subtype in EBV-associated Nasopharyngeal Carcinoma from Vietnam

Abstract

The Epstein-Barr nuclear antigen 1 (EBNA-1) gene, plays a key role in viral infection, immortalization, viral genome replication, transcription and maintenance, and is the frequently detected gene, protein in both latent and lytic stage of Epstein-Barr virus (EBV). Based on the amino acid at position 487, EBNA-1 was classified into five subtypes, including P-Ala, P-Thr, V-Val, V-Pro and V-Leu. In Vietnam, an Asian country with a high incidence, mortality rates of nasopharyngeal carcinoma (NPC), had limited research on the EBNA-1 variation. Therefore, the aim of the current study was to identify the pattern of the EBNA-1 V-Val subtype in Vietnamese NPC patients, for its value further applied in NPC patients. Fifty-eight NPC biopsy samples were collected from local patients, analyzed by nested-polymerase chain reaction (nested-PCR), sequencing and compared to a previous B95-8 prototype sequence. Four EBNA-1 subtypes, including V-Val (35/44, 79.55%), P-Ala (2/44, 4.55%), P-Thr (5/44, 11.36%), and V-Leu (2/44, 4.55%), were observed in 44/58 samples. The sequences of the V-Val subtype were compared to the B95-8 prototype, resulting in five patterns, contained seven consensus changes, including five amino acid changes at positions 487, 499, 502, 524, 594, and two silent changes at residues 520 and 553. Of these, four of five, patterns were identified as novel patterns of the V-Val subtype, showing the different changes of amino acids at positions 492, 528, 529, 553, 585 and 588, by comparison with previous studies of V-Val EBNA-1. Those data suggested the profile of variation patterns of the EBNA-1 gene, related to geographic distribution, in Vietnamese NPC patients.

Keywords: Epstein-Barr nuclear antigen (EBNA-1); Epstein-Barr virus (EBV); V-val; Variation.

Figure 1

Schematic of the EBNA-1 gene and position of primers aligned with B95-8 strain (V10555). Primer 1: EBNA-1-1; primer 2: EBNA-1-2; primer 3: EBNA-1-3 and primer 4: EBNA-1-4.

Figure 2

Electrophoresis of nested-PCR products of the represented NPC biopsy samples. T91, T92, T93, T94, T95, T96: clinical biopsy samples; N: negative control; MW: molecular weight 100 bp.

Figure 3

Represented sequence identification of EBNA-1 subtype based on the amino acid at position 487. A) T91: V-Val subtype; B) V-Leu subtype; C) P-Thr subtype; D) P-Ala subtype.

Figure 4

Represented nucleotide substitution consensus changes indicated by T5. Amino acid at position: A) codon 497; B) codon 499; C) codon 502; D) codon 520; E) codon 524; F) codon 553; G) codon 594.

Figure 5

Represented nucleotide/amino acid additional changes: A) codon 585 by T6; B) codons 528 and 529 by T77; C) codon 588 by T77; D) codon 492 by T75.