Epstein-Barr virus protein EB2 contains an N-terminal transferable nuclear export signal that promotes nucleocytoplasmic export by directly binding TAP/NXF1.

The Epstein-Barr virus early protein EB2 (also called BMLF1, Mta, or SM), which allows the nuclear export of a subset of early and late viral mRNAs derived from intronless genes, is essential for the production of infectious virions. An important feature of mRNA export factors is their capacity to shuttle continuously between the nucleus and the cytoplasm. In a previous study, we identified a novel CRM1-independent transferable nuclear export signal (NES) at the N terminus of EB2, between amino acids 61 and 146. Here we show that this NES contains several small arginine-rich domains that cooperate to allow efficient interaction with TAP/NXF1. Recruitment of TAP/NXF1 correlates with this NES-mediated efficient nuclear export when it is fused to a heterologous protein. Moreover, the NES can export mRNAs bearing MS2 RNA-binding sites from the nucleus when tethered to the RNA via the MS2 phage coat protein RNA-binding domain.

Protein kinase CK2 phosphorylation of EB2 regulates its function in the production of Epstein-Barr virus infectious viral particles.

The Epstein-Barr Virus (EBV) early protein EB2 (also called BMLF1, Mta, or SM) promotes the nuclear export of a subset of early and late viral mRNAs and is essential for the production of infectious virions. We show here that in vitro, protein kinase CK2alpha and -beta subunits bind both individually and, more efficiently, as a complex to the EB2 N terminus and that the CK2beta regulatory subunit also interacts with the EB2 C terminus. Immunoprecipitated EB2 has CK2 activity that phosphorylates several sites within the 80 N-terminal amino acids of EB2, including Ser-55, -56, and -57, which are localized next to the nuclear export signal. EB2S3E, the phosphorylation-mimicking mutant of EB2 at these three serines, but not the phosphorylation ablation mutant EB2S3A, efficiently rescued the production of infectious EBV particles by HEK293(BMLF1-KO) cells harboring an EB2-defective EBV genome. The defect of EB2S3A in transcomplementing 293(BMLF1-KO) cells was not due to impaired nucleocytoplasmic shuttling of the mutated protein but was associated with a decrease in the cytoplasmic accumulation of several late viral mRNAs. Thus, EB2-mediated production of infectious EBV virions is regulated by CK2 phosphorylation at one or more of the serine residues Ser-55, -56, and -57.