Cell phenotype-dependent splicing reflecting differential promoter usage for EBNA transcripts in EBV-carrying cells

Area of Science:

• Virology
• Molecular Biology
• Oncology

Background:

• Epstein-Barr virus (EBV) exhibits varied latent infection patterns in different host cells.
• Three EBV latency types are defined by the expression of EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs).
• Specific EBV promoters, including Qp, Cp, and Wp, are implicated in EBNA transcription.

Purpose of the Study:

• To investigate the alternative EBNA mRNA transcription programs utilized by EBV in different cellular contexts.
• To elucidate the role of cellular phenotype in regulating EBV promoter usage.
• To understand the switch in EBV transcription programs during cellular differentiation or hybridization.

Main Methods:

• RNA-reverse transcription polymerase chain reaction (RT-PCR) was employed to analyze EBNA mRNA splicing and transcription initiation sites.
• Somatic cell hybridization was used to downregulate EBNA 2-6 expression and observe changes in promoter usage.
• Analysis of EBV promoter usage in various cell lines including Burkitt's lymphoma (BL), nasopharyngeal carcinoma (NPC), and lymphoblastoid cell lines (LCL).

Main Results:

• EBV-infected cells lacking EBNA 2-6 expression splice EBNA mRNA at the Q-exon, indicating Q-promoter activation.
• Cells expressing EBNA 2-6 utilize upstream promoters (Cp or Wp) for EBNA transcription.
• Somatic cell hybridization with non-B cells or NPC cells induced Q-promoter usage, mirroring the 'EBNA-1 only' program.

Conclusions:

• EBV employs at least two distinct transcription programs, regulated by the host cell phenotype.
• The Q-promoter-driven 'EBNA-1 only' program is activated in non-B cell phenotypes.
• Upstream promoters (Cp/Wp) generate a long pre-mRNA for all EBNAs in immunoblastic cells (LCL, BL group III).