Functional interaction of nuclear factor y and sp1 is required for activation of the epstein-barr virus C promoter.
Two Epstein-Barr virus (EBV) latent cycle promoters, Wp and Cp, are activated sequentially during virus-induced transformation of primary B lymphocytes. Immediately postinfection, viral transcription initiates from Wp, leading to expression of EBV nuclear antigen 2 (EBNA2) and EBNA5. Within 36 h, there is a switch in promoter usage from Wp to the upstream Cp, which leads to expression of EBNA1 to EBNA6. EBNA2 appears to be required for the Wp-to-Cp switch, but the switching mechanism is not fully understood at the molecular level. In a previous investigation we showed that there is an EBNA2-independent activity of reporter constructs containing deletion fragments of Cp in B-lymphoid cell lines, and we demonstrated that Cp activity is highly dependent on several cellular transcription factors, including nuclear factor Y (NF-Y) and Sp1. In the present work, we analyzed the effect of NF-Y on Cp activity in greater detail. We demonstrate that (i) a dominant negative analogue of NF-Y abolishes Cp activity, (ii) NF-Y and Sp1 costimulate Cp, and (iii) the oriPI-EBNA1-induced transactivation of Cp requires concomitant expression of NF-Y and Sp1, although additional factors seem necessary for optimal activation. Furthermore, using the lymphoblastoid cell line EREB2-5, in which EBNA2 function is regulated by estrogen, we demonstrate that inactivation of EBNA2 results in decreased expression of NF-Y and down-regulation of Cp. On reconstitution of the EBNA2 function, the cells enter the cell cycle, NF-Y levels increase, and a concomitant Wp-to-Cp switch occurs. Taken together, our results suggest that NF-Y is essential for Cp activation and that up-regulation of NF-Y may contribute to a successful Wp-to-Cp switch during B-cell transformation.
Boreström
Cecilia
C
Department of Clinical Chemistry and Transfusion Medicine, Sahlgrenska University Hospital, Göteborg University, S-413 45 Gothenburg, Sweden.
Zetterberg
Henrik
H
Liff
Kristian
K
Rymo
Lars
L
D001402:B-Lymphocytes / Q000166:cytology / Q000378:metabolism; D023081:CCAAT-Binding Factor / Q000378:metabolism*; D002455:Cell Division; D002461:Cell Line, Transformed; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
D023081:CCAAT-Binding Factor; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; C402244:nuclear factor Y