B cells infected with Type 2 Epstein-Barr virus (EBV) have increased NFATc1/NFATc2 activity and enhanced lytic gene expression in comparison to Type 1 EBV infection

• Published in: PLoS pathogens 2020-02, Vol.16 (2), p.e1008365-e1008365
• Main Authors: Romero-Masters, James C, Huebner, Shane M, Ohashi, Makoto, Bristol, Jillian A, Benner, Bayleigh E, Barlow, Elizabeth A, Turk, Gail L, Nelson, Scott E, Baiu, Dana C, Van Sciver, Nicholas, Ranheim, Erik A, Gumperz, Jenny, Sherer, Nathan M, Farrell, Paul J, Johannsen, Eric C, Kenney, Shannon C
• Format: Article
• Language: English
• Subjects: Analysis; Animals; B cells; B-cell lymphoma; B-cell receptor; B-Lymphocytes - metabolism; B-Lymphocytes - virology; Binding sites; Biology and Life Sciences; BZLF1 protein; Ca2+/calmodulin-dependent protein kinase IV; Calcium; Calcium signalling; Cell Line; Cord blood; Cyclosporins; DNA binding proteins; DNA-Binding Proteins - metabolism; Epstein-Barr virus; Epstein-Barr Virus Infections - genetics; Epstein-Barr Virus Nuclear Antigens; Gene expression; Gene Expression - genetics; Gene Expression Regulation, Viral - genetics; Genes; Genomes; Health aspects; Herpesvirus 4, Human - genetics; Herpesvirus 4, Human - metabolism; Herpesvirus 4, Human - pathogenicity; Humans; Immunology; Infections; Latency; Lymphoblastoid cell lines; Lymphocytes B; Lymphoma; Lymphomas; Medicine; Medicine and Health Sciences; Mice; NF-AT protein; NFATC Transcription Factors - genetics; Oncology; Phenotypes; Promoter Regions, Genetic - genetics; Public health; Research and Analysis Methods; Signaling; Strains (organisms); Trans-Activators - genetics; Trans-Activators - metabolism; Transcription factors; Transcription Factors - metabolism; Tumors; Viral infections; Viral Proteins - metabolism; Virus Activation; Virus Latency; Viruses
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1008365

Humans are infected with two distinct strains (Type 1 (T1) and Type 2 (T2)) of Epstein-Barr virus (EBV) that differ substantially in their EBNA2 and EBNA 3A/B/C latency genes and the ability to transform B cells in vitro. While most T1 EBV strains contain the "prototype" form of the BZLF1 immediate-early promoter ("Zp-P"), all T2 strains contain the "Zp-V3" variant, which contains an NFAT binding motif and is activated much more strongly by B-cell receptor signalling. Whether B cells infected with T2 EBV are more lytic than cells infected with T1 EBV is unknown. Here we show that B cells infected with T2 EBV strains (AG876 and BL5) have much more lytic protein expression compared to B cells infected with T1 EBV strains (M81, Akata, and Mutu) in both a cord blood-humanized (CBH) mouse model and EBV-transformed lymphoblastoid cell lines (LCLs). Although T2 LCLs grow more slowly than T1 LCLs, both EBV types induce B-cell lymphomas in CBH mice. T1 EBV strains (M81 and Akata) containing Zp-V3 are less lytic than T2 EBV strains, suggesting that Zp-V3 is not sufficient to confer a lytic phenotype. Instead, we find that T2 LCLs express much higher levels of activated NFATc1 and NFATc2, and that cyclosporine (an NFAT inhibitor) and knockdown of NFATc2 attenuate constitutive lytic infection in T2 LCLs. Both NFATc1 and NFATc2 induce lytic EBV gene expression when combined with activated CAMKIV (which is activated by calcium signaling and activates MEF2D) in Burkitt Akata cells. Together, these results suggest that B cells infected with T2 EBV are more lytic due to increased activity of the cellular NFATc1/c2 transcription factors in addition to the universal presence of the Zp-V3 form of BZLF1 promoter.