Epstein-Barr virus recombinants with specifically mutated BCRF1 genes

Abstract

Epstein-Barr virus (EBV) recombinants with specifically mutated BCRF1 genes were constructed and compared with wild-type BCRF1 recombinants derived in parallel for the ability to initiate and maintain latent infection and growth transformation in primary human B lymphocytes. A stop codon insertion after codon 116 of the 170-codon BCRF1 open reading frame or deletion of the entire gene had no effect on latent infection, B-lymphocyte proliferation into long-term lymphoblastoid cell lines (LCLs), or virus replication. LCLs infected with the stop codon recombinant were indistinguishable from wild-type recombinant-infected LCLs in tumorigenicity in SCID mice. However, mutant BCRF1 recombinant-infected cells differed from wild-type recombinant-infected cells in their inability to block gamma interferon release in cultures of permissively infected LCLs incubated with autologous human peripheral blood mononuclear cells. This is the first functional assay for BCRF1 expression from the EBV genome. BCRF1 probably plays a key role in modulating the specific and nonspecific host responses to EBV infection.

Reducing the complexity of the transforming Epstein-Barr virus genome to 64 kilobase pairs

“…TRANSFORMING MINI-EBV GENOMES 991 40,41). Together with the previous experiments, the current experiments demonstrate that the unique EBV DNA required for B-lymphocyte transformation is less than 64 kbp.…”

“…Incorporation of small segments of homologous DNAs from the endogenous P3HR-1 genome occurred in these experiments, although it was infrequent. Nonhomologous recombination was unusual, as in previous experiments of this type (33,(40)(41)(42)(43)(44)(45). From the results in these and previous experiments, recombination with the endogenous P3HR-1 genome is unlikely to occur near sites of nonhomology, such as pertains when the transfected DNA is deleted for a large segment (33).…”

“…Recombination among the three transfected cosmids results in a 46-kbp deletion relative to the B95-8 EBV genome and a 58-kbp deletion relative to wild-type (WT) EBV genomes. Transforming EBV recombinants have also been constructed which are deleted for the LMP2 (bp 02 to 648), EBERs (bp 6612 to 7263), BCRF1 (bp 9535 to 12870), and BHRF1 (bp 53721 to 60612) encoding DNA segments (23,26,40,41).…”

Virus Attenuation after Deletion of the Cytomegalovirus Fc Receptor Gene Is Not due to Antibody Control

“…The fcr-1 gene is nonessential for replication in cell culture and has no phenotype in vitro. Herpesvirus functions have been shown to modulate virus-host interactions such as antigen presentation and immune system control (2,4,12,15,17,31,34,37,(39)(40)(41), tissue tropism (3-5, 23, 27), virus spread (9,25,42), and the establishment of latency (33). Most of these functions are encoded by nonessential genes.…”

“…DNA viruses have developed numerous strategies to modulate or evade the immune system control of the host. Immune system modulators encoded within viral genomes include proteins that interrupt the complement cascade, act as cytokines or cytokine antagonists (34,37,40), inhibit the effector mechanisms mediated through antibodies, or interfere with antigen processing and presentation pathways (reviewed in references 15 and 28). Cytomegaloviruses (CMV) genomes encode several genes whose products modify the efficiency of host immune system control.…”