Epstein-Barr virus nuclear protein 3A domains essential for growth of lymphoblasts: transcriptional regulation through RBP-Jkappa/CBF1 is critical.

Experimental reverse genetic replacement of Epstein-Barr virus nuclear antigen 3A (EBNA3A) with a conditional mutant EBNA3A revealed that EBNA3A is critical for continued lymphoblastoid cell (LCL) growth. Wild-type (wt) EBNA3A expressed in the LCLs specifically sustained growth under nonpermissive conditions, whereas EBNA3B or EBNA3C expression had no effect (S. Mauro, E. Johannsen, D. Illanes, A. Cooper, and E. Kieff, J. Virol. 77:10437-10447, 2003). This genetic system and related biochemical assays have now been used to discover that EBNA3A lacking amino acid residues 170 to 240 (delta170-240), TLGC202 to AAGA202, or delta300-386, which are deficient in repression of EBNA2 activation of an RBP-Jkappa/CBF1-dependent EBV Cp enhancer, are null mutations for LCL growth, whereas EBNA3A delta2-124, delta410-439, delta440-470, delta470-500, delta500-523, delta523-612, and delta620-820, which are wt in repression are wt for LCL growth. Thus, EBNA3A regulation of transcription through RBP-Jkappa/CBF1 is critical for LCL growth. EBNA3A mutants deleted of amino acid residues 240 to 300, 386 to 410, or 827 to 944 were intermediate, null, or intermediate, respectively, for LCL growth despite being wt for RBP-Jkappa association and repression. Amino acid residues 240 to 300, 386 to 410, and, particularly, C-terminal residues 827 to 944 are therefore likely to contribute to LCL growth through RBP-Jkappa-independent mechanisms.