Characterization of tau antigens isolated from uninfected and simian virus 40-infected monkey cells and papovavirus-transformed cells.

Tau antigens (also known as cellular or nonviral tumor antigens) were detected in uninfected and simian virus 40-infected monkey cells after immunoprecipitation with serum from hamsters bearing simian virus 40-induced tumours (anti-T serum). These two proteins (56,000 daltons) were digested to similarly sized peptides with various amounts of Staphylococcus aureus V8 protease. The Tau antigen isolated from infected monkey cells was closely related but was not identical to the corresponding protein from human cells transformed by simian virus 40, as determined by two-dimensional mapping of their methionine-labeled tryptic peptides. Hamster cells transformed by various primate papovaviruses (simian virus 40, BK virus, and JC virus) synthesized indistinguishable Tau antigens, as determined by two-dimensional peptide mapping. When tested by the same procedure, these proteins and the ones made in monkey and human cells were found to be related to the Tau antigens isolated from simian virus 40-transformed mouse and rat cells. Based on these results, an "evolutionary tree" was constructed to show the relationship among the methionine-containing tryptic peptides of all of these proteins.     

Novel Tat-encoding bicistronic human immunodeficiency virus type 1-based gene transfer vectors for high-level transgene expression

We describe bicistronic single-exon Tat (72-amino-acid Tat [Tat72])- and full-length Tat (Tat86)-encoding gene transfer vectors based on human immunodeficiency virus type 1 (HIV-1). We created versions of these vectors that were rendered Rev independent by using the constitutive transport element (CTE) from Mason-Pfizer monkey virus (MPMV). Tat72-encoding vectors performed better than Tat86-expressing vectors in gene transfer experiments. CTE-containing vectors, produced in a Rev-independent packaging system, had gene transfer efficiencies nearly equivalent to those produced using a combination RNA transport (CTE and Rev-Rev response element)-based packaging system. The Tat72-encoding vectors could be efficiently transduced into a variety of cell types, showed higher levels of transgene expression than vectors with the simian cytomegalovirus immediate-early or the simian virus 40 early promoter, and provide an alternative to HIV-1 vectors with internal promoters.     

Analysis of an activatable promoter: sequences in the simian virus 40 late promoter required for T-antigen-mediated trans activation.

The late promoter of simian virus 40 (SV40) is activated in trans by the viral early gene product, T antigen. We inserted the wild-type late-promoter region, and deletion mutants of it, into chloramphenicol acetyltransferase transient expression vectors to identify promoter sequences which are active in the presence of T antigen. We defined two promoter activities. One activity was mediated by a promoter element within simian virus 40 nucleotides 200 to 270. The activity of this element was detectable only in the presence of an intact, functioning origin of replication and accounted for 25 to 35% of the wild-type late-promoter activity in the presence of T antigen. The other activity was mediated by an element located within a 33-base-pair sequence (simian virus nucleotides 168 to 200) which spans the junction of the 72-base-pair repeats. This element functioned in the absence of both the origin of replication and the T-antigen-binding sites and appeared to be responsible for trans-activated gene expression. When inserted into an essentially promoterless plasmid, the 33-base-pair element functioned in an orientation-dependent manner. Under wild-type conditions in the presence of T antigen, the activity of this element accounted for 65 to 75% of the late-promoter activity. The roles of the 33-base-pair element and T antigen in trans-activation are discussed.