Viral Late Promoter Research Articles

Sequences downstream of the RNA initiation site regulate human T-cell lymphotropic virus type I basal gene expression.

Sequences which control basal human T-cell lymphotropic virus type I (HTLV-I) transcription probably play an important role in initiation and maintenance of virus replication. We have identified and analyzed a 45-nucleotide sequence (downstream regulatory element 1 [DRE 1]) at the boundary of the R/U5 region of the long terminal repeat which is required for HTLV-I basal transcription. The basal promoter strength of constructs that contained deletions in the R/U5 region of the HTLV-I long terminal repeat were analyzed by chloramphenicol acetyltransferase assays following transfection of Jurkat T cells. We consistently observed a 10-fold decrease in basal promoter activity when sequences between +202 to +246 were deleted. By reverse transcriptase polymerase chain reaction RNA analysis, we confirmed that the drop in chloramphenicol acetyltransferase activity was paralleled by a decrease in the level of steady-state RNA. DRE 1 did not affect the level of Tax1 transactivation. Using a gel shift assay, we have purified a highly enriched fraction that could specifically bind DRE 1. This DNA affinity column fraction contained four detectable proteins on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis: p37, p50, p60, and p100. The affinity column fraction stimulated HTLV-I transcription approximately 12-fold in vitro. No effect was observed with the human immunodeficiency virus or adenovirus major late promoters. Following renaturation of the proteins isolated from an SDS-containing gel, p37, but not the other protein fractions, was able to specifically bind to DRE 1.

Self-assembly of human papillomavirus type 1 capsids by expression of the L1 protein alone or by coexpression of the L1 and L2 capsid proteins

Vaccinia virus vectors were used to express the major (L1) and minor (L2) capsid proteins of human papillomavirus type 1 (HPV-1) with the vaccinia virus early (p7.5K) or late (pSynth, p11K) promoters. All constructs expressed the appropriate-sized HPV proteins, and both L1 and L2, singly or in combination, localized to the nucleus. Capsids were purified by cesium chloride density gradient centrifugation from nuclei of cells infected with a vaccinia virus-L1 (vac-L1) recombinant or a vac-L1-L2 recombinant but not from vac-L2-infected cells. Electron microscopy showed that the particles were 55 nm in diameter and had icosahedral symmetry. Immunogold-labeled antibodies confirmed the presence of the L1 and L2 proteins in the HPV-1 capsids. Capsids containing L1 alone were fewer and more variable in size and shape than capsids containing the L1 and L2 proteins. The L1-plus-L2 capsids were indistinguishable in appearance from HPV-1 virions obtained from plantar warts. The ability to produce HPV capsids in vitro will be useful in many studies of HPV pathogenicity.

Evidence that a sequence similar to TAR is important for induction of the JC virus late promoter by human immunodeficiency virus type 1 Tat.

A specific RNA sequence located in the leader of all human immunodeficiency virus type 1 (HIV-1) mRNAs termed the transactivation response element, or TAR, is a primary target for induction of HIV-1 long terminal repeat activity by the HIV-1-derived trans-regulatory protein, Tat. Human neurotropic virus, JC virus (JCV), a causative agent of the degenerative demyelinating disease progressive multifocal leukoencephalopathy, contains sequences in the 5' end of the late RNA species with an extensive homology to HIV-1 TAR. In this study, we examined the possible role of the JCV-derived TAR-homologous sequence in Tat-mediated activation of the JCV late promoter (Tada et al., Proc. Natl. Acad. Sci. USA 87:3479-3483, 1990). Results from site-directed mutagenesis revealed that critical G residues required for the function of HIV-1 TAR that are conserved in the JCV TAR homolog play an important role in Tat activation of the JCV promoter. In addition, in vivo competition studies suggest that shared regulatory components mediate Tat activation of the JCV late and HIV-1 long terminal repeat promoters. Furthermore, we showed that the JCV-derived TAR sequence behaves in the same way as HIV-1 TAR in response to two distinct Tat mutants, one of which that has no ability to bind to HIV-1 TAR and another that lacks transcriptional activity on a responsive promoter. These results suggest that the TAR homolog of the JCV late promoter is responsive to HIV-1 Tat induction and thus may participate in the overall activation of the JCV late promoter mediated by this transactivation.

Nonreplicating vaccinia vector efficiently expresses recombinant genes.

Modified vaccinia Ankara (MVA), a highly attenuated vaccinia virus strain that has been safety tested in humans, was evaluated for use as an expression vector. MVA has multiple genomic deletions and is severely host cell restricted: it grows well in avian cells but is unable to multiply in human and most other mammalian cells tested. Nevertheless, we found that replication of viral DNA appeared normal and that both early and late viral proteins were synthesized in human cells. Proteolytic processing of viral structural proteins was inhibited, however, and only immature virus particles were detected by electron microscopy. We constructed an insertion plasmid with the Escherichia coli lacZ gene under the control of the vaccinia virus late promoter P11, flanked by sequences of MVA DNA, to allow homologous recombination at the site of a naturally occurring 3500-base-pair deletion within the MVA genome. MVA recombinants were isolated and propagated in permissive avian cells and shown to express the enzyme beta-galactosidase upon infection of nonpermissive human cells. The amount of enzyme made was similar to that produced by a recombinant of vaccinia virus strain Western Reserve, which also had the lacZ gene under control of the P11 promoter, but multiplied to high titers. Since recombinant gene expression is unimpaired in nonpermissive human cells, MVA may serve as a highly efficient and exceptionally safe vector.

A cellular promoter-based expression cassette for generating recombinant baculoviruses directing rapid expression of passenger genes in infected insects

We have developed an expression cassette which allows the generation of recombinant baculoviruses that can express passenger genes under the control of a constitutive cellular promoter derived from the cytoplasmic actin gene of the silkmoth Bombyx mori. Silkmoth tissue culture cells which were infected with a recombinant B. mori nuclear polyhedrosis virus (BmNPV) containing the gene-encoding chloramphenicol acetyl transferase (CAT) under the control of this expression cassette expressed significant CAT activity beginning 5 hr postinfection (p.i.). Cells infected with a recombinant BmNPV containing the cat gene under the control of the polyhedrin gene promoter did not express CAT activity until 20 hr p.i. Silkworm larvae were also infected with the two recombinant viruses by hemocelic injections and all larval tissues examined were found to express the cat gene. While significant actin-cassette-driven CAT expression in vivo was first seen at 24 hr p.i., expression from the polyhedrin promoter was not seen until 48 hr p.i. By 60 hr p.i., tissues of larvae infected with the recombinant virus expressing cat under polyhedrin promoter control were found to exhibit sixfold higher CAT activity than those infected with recombinant virus expressing the cat gene under the control of the actin promoter. The 24-hr temporal advantage in expression of a passenger gene in infected larvae indicates that the actin-promoter-based expression cassette or other analogous cellular promoter-based cassettes could be used for generating recombinant baculovirus insecticides which could incapacitate pest insects more quickly than viruses employing the polyhedrin or other late viral promoters for expressing insect-incapacitating proteins.

Regulated expression of foreign genes in vaccinia virus under the control of bacteriophage T7 RNA polymerase and the Escherichia coli lac repressor.

The gene encoding bacteriophage T7 RNA polymerase (T7gene1) was placed under the control of regulatory elements from the Escherichia coli lac operon to construct an inducible vaccinia virus expression system consisting entirely of prokaryotic transcriptional machinery. Regulated expression of T7 RNA polymerase was necessary to construct a stable recombinant vaccinia virus harboring a T7 promoter; otherwise, uncontrolled expression led to interference with endogenous virus replication. To this end, the gene encoding the repressor protein of the lac operon was fused to a viral early/late promoter so that it was expressed constitutively, and the lac operator was interposed between a viral major late promoter and T7gene1. Greater than 99% repression of T7 RNA polymerase, which was relieved approximately 80-fold in the presence of the inducer isopropyl-beta-D-thiogalactopyranoside (IPTG), was obtained. An expression cassette containing a T7 promoter-controlled beta-galactosidase reporter gene was recombined into a different region of the viral genome containing T7gene1. A stable, double recombinant virus was isolated and grown to a high titer. In the absence of inducer, beta-galactosidase expression was substantially repressed. Addition of increasing amounts of IPTG induced expression of beta-galactosidase to the point of suppression of viral replication. This hybrid vaccinia virus system (Vac/Op/T7) has potential applications for the efficient bioproduction of a wide variety of gene products.

Expression of a Paralytic Neurotoxin Gene to Improve Insect Baculoviruses as Biopesticides

Recombinant baculoviruses expressing tox34, a mite gene encoding the insect–selective neurotoxin TxP–I, paralyze insects during the infection process. In this study, we investigated several key parameters which can influence the relative efficacy of toxin–expressing recombinant baculoviruses as biological pesticides. These parameters included the nature of the promoter controlling tox34 transcription and polyhedra production in infected larvae. The amount of weight gained during infection and the time required for paralysis was promoter dependent; the most effective promoter tested was a hybrid comprising both late and very late viral promoter elements. Since TxP–I paralyzes but does not immediately kill infected insects, larvae can be used for virus pesticide production.

The ICP4 binding sites in the herpes simplex virus type 1 glycoprotein D (gD) promoter are not essential for efficient gD transcription during virus infection.

Activation of the early and late genes of herpes simplex virus type 1 during infection in tissue culture requires functional immediate-early regulatory protein ICP4. ICP4 is a specific DNA-binding protein which recognizes a variety of DNA sequences, many of which contain the consensus ATCGTC. In general, mutations which impair the ability of ICP4 to bind to DNA also eliminate its ability to activate viral early and late promoters both in transfection assays and in the infected cell. However, the role of ICP4 binding sites in the viral genome is unclear; many early and late promoters do not contain consensus binding sites in their vicinity. The glycoprotein D (gD) gene contains two well-characterized ICP4 binding sites upstream of its promoter and a third downstream of the transcription start site. Multimerization of one of these sites has been shown to increase the response of the gD promoter to ICP4 in transfection assays, while their removal reduces stimulation of the gD promoter by ICP4 in vitro. To assess the role of these binding sites during virus infection, we have constructed a recombinant viral genome which has mutations affecting all three. Comparison of the amounts of gD RNA synthesized by the recombinant and wild-type viruses indicated that the mutations had little or no effect on the activity of the gD promoter. Therefore, either the sites have no essential role in gD promoter regulation in the presence of all of the herpes simplex virus type 1 IE polypeptides during a normal infection or they can be functionally substituted by other ICP4 binding sites elsewhere in the genome.

Adenovirus DNA replication facilitates binding of the MLTF/USF transcription factor to the viral major late promoter within infected cells.

The activity of the adenovirus major late promoter is substantially increased as the infection proceeds from the early to late phase. To gain insight into the regulation of this promoter, we analyzed protein-DNA interactions by in vivo DMS and DNasel footprinting during the course of adenovirus infection. Little or no protein interaction at promoter sequences was detected early (5 hr) after infection but strong interactions at the major late transcription factor (MLTF/USF) binding site and at the TATA box were evident late (12 hr) after infection. Comparison of in vivo and in vitro footprints revealed that the in vivo interaction late after infection results from binding of the cellular transcription factor MLTF/USF. Nuclear extracts prepared from uninfected cells as well as cells harvested at 5 and 12 hr after infection contained similar levels of MLTF/USF footprint activity, therefore the lack of a detectable interaction early after infection is not due to reduced levels of the factor early in the viral growth cycle. Viral DNA replication was required for MLTF/USF binding at the major late promoter. These results indicate that DNA replication participates in the regulation of adenovirus late gene expression by facilitating the binding of a transcription factor to the major late promoter.

Identification of a novel transcription factor, ACF, in cultured avian fibroblast cells that interacts with a Marek's disease virus late gene promoter

Interactions between factors in duck and chick embryo fibroblast (DEF and CEF, respectively) nuclear extracts and the Marek's disease virus (MDV) gp57–65 gene promoter were investigated. Results of in vitro transcription and gel mobility-shift assays indicated that multiple cellular factors interact with 5′-flanking sequences of the MDV gp57–65 gene. One sequence-specific DNA binding activity (termed ACF for avian cell factor(s)) was identified by interaction of DEF and CEF nuclear extract proteins with a particular site (nucleotides −193 to −177) in the MDV gp57–65 gene promoter. Binding of ACF to its apparent recognition sequence, contained within the 17-bp oligonucleotide 5′-CTAGTTTACTTGTTTGT-3′ (ACF-12), was highly sequence-specific. Radiolabeled ACF-12 oligonucleotide bound significant ACF protein in the presence of a 400-fold molar excess of unlabeled nonspecific competitor DNA. A similar amount of specific competitor completely abolished ACF binding to probe DNA. Deletion of the ACF binding site from MDV gp57–65 gene promoters linked to a chloramphenicol acetyltransferase (CAT) reporter gene reduced expression of CAT activity by twofold relative to that seen with a gp57–65 promoter-CAT construct containing an intact ACF binding site. Transfection inhibition assays using double-stranded ACF binding site competitors reduced steady-state levels of gp57–65 mRNA in MDV infected cells by over twofold relative to those in control infected cells. Introduction of a similar amount of nonspecific double-stranded oligonucleotide had no adverse effect on gp57–65 mRNA levels. These data suggest that ACF is important for efficient expression of gp57–65.

Characterization of a potent varicella-zoster virus-encoded trans-repressor.

Using a transient expression assay in Vero cells, we have shown that the protein product from gene 61 of varicella-zoster virus (VZV) can repress the function of the VZV encoded trans-activators on putative viral immediate-early, early, and late gene promoters. The repression is exerted at the transcriptional level and requires functional gene 61 protein. This trans-repressor is the herpes simplex type 1 ICP0 (a trans-activator) homolog, as defined by gene location, the sharing of a cysteine-rich putative zinc-binding finger in the amino-terminal region, and limited amino acid homology. Open reading frame 61 (ORF61)-mediated trans-repression appears to be specific for VZV-encoded trans-activators in that it has no effect on simian virus 40 and Rous sarcoma virus promoters. Moreover, it does not inhibit trans-activation of the human T-lymphotropic virus type I and human immunodeficiency virus long terminal repeats by tax and tat genes, respectively. We constructed plasmids with mutations in ORF61 and tested them for their ability to inhibit trans-activator (VZV genes 4 and 62)-mediated activation of the viral thymidine kinase promoter-chloramphenicol acetyltransferase construct. Mutants containing interruptions in ORF61 lost their trans-repressing ability, as demonstrated at both the protein and steady-state RNA levels. These results suggest that the ORF61 protein product can mediate down-regulation of VZV gene expression.

A recombinant cDNA derived from human brain encodes a DNA binding protein that stimulates transcription of the human neurotropic virus JCV

The human neurotropic virus JCV contains a 98-base pair repeat enhancer/promoter sequence that confers glial-specific transcription to the viral early and late promoters. The central region of this repeat, designated the B-domain, binds to a glial-derived nuclear protein that stimulates transcription of the viral promotor in vitro. We now report the isolation of a recombinant cDNA clone, termed glial factor-1 (GF1), from a brain expression library that encodes a novel protein which interacts with the JCV B-domain. Results from RNA studies indicate that the GF1 transcript is more abundant in brain than in other tissues and that the level of GF1 RNAs increases progressively during brain development. Cotransfection of the recombinant GF1 expressor plasmid with JCV promoters indicates that GF1 stimulates transcription of the JCV late promoter and to a lesser extent the JCV early promoter predominantly in cells of human glial origin. Thus, GF1 is a sequence-specific DNA binding protein that may play a role in determining the glial-specific expression of JCV.

Endogenously produced interferon alpha protects mice from herpes simplex virus type 1 corneal disease.

Intravenous (i.v.) injection of u.v. light-inactivated herpes simplex virus type 1 (UV HSV-1) at the time of HSV-1 corneal infection reduced the cytotoxic T lymphocyte (CTL) response to HSV-1, and significantly reduced the incidence of HSV-1-induced corneal stromal disease in A/J mice. The spread of HSV-1 through the eye after corneal infection, detected using engineered HSV-1 (US3::Tn5-lacZ) with the lacZ gene under the transcriptional control of the viral late gene promoter for glycoprotein C, was also markedly reduced by i.v. UV HSV-1 injection. The restriction of HSV-1 corneal invasiveness in i.v. UV HSV-1-injected mice preceded the onset of a detectable specific cell-mediated or humoral immune response to HSV-1, and was accompanied by an elevated serum titre of interferon (IFN-alpha), reversed by anti-IFN-alpha/beta antibody, and mimicked by systemic IFN-alpha treatment. IFN-alpha-treated mice developed a normal CTL response to HSV-1 after corneal infection, but the corneal invasiveness of the virus was markedly reduced and none of the treated mice developed corneal stromal disease. Together with our previous findings that HSV-1-specific CTLs participate in the pathogenesis of corneal stromal disease, these results indicate that i.v. injection of UV HSV-1 at the time of corneal infection may prevent stromal disease by the combined effects of IFN-mediated reduction of the spread of virus in the cornea and inhibition of the activity of the HSV-specific T lymphocytes that induce tissue destruction in the corneal stroma.

Transcription activation at a distance by phage φ29 protein p4: Effect of bent and non-bent intervening DNA sequences

Protein p4 of the Bacillus subtilis phage φ29 switches on the transcription of the viral late genes by binding to the viral late promoter at a region close to the RNA polymerase binding site. Gel retardation and DNase I footprinting assays show that the presence of protein p4 is required for RNA polymerase recognition of the late promoter. The protein p4 and RNA polymerase DNA binding sites have been separated by the insertion of bent and non-bent DNA sequences of different lengths. These mutant promoters were used to study in vitro their protein p4-dependent transcriptional activity and their interaction with both protein p4 and RNA polymerase. The results indicate that protein p4 is able to function at longer DNA distances from the RNA polymerase binding site than in the natural promoter. The extent of protein p4 activity depended on the length and conformation of the inserted DNA. Activation of transcription and RNA polymerase binding was favoured when the relative orientation of protein p4 and RNA polymerase was conserved and when the intervening DNA had a bent conformation. These data, together with the DNase I footprints, suggest that activation at distance by protein p4 involves a DNA loop held by the interaction of protein p4 and RNA polymerase.

The induction of cytotoxic T-lymphocyte precursor cells by recombinant vaccinia virus expressing human papillomavirus type 16 L1

Expression of the major coat protein L1 of human papillomavirus type 16 by recombinant vaccinia viruses using a vaccinia late promoter and their use in generating antibodies have already been reported (Zhou et al., 1990). We have now constructed recombinant vaccinia viruses (VVs) which express the Li protein from an early promoter with the intention of inducing cell-mediated immunity. This necessitated the removal of sequence motifs (TTTTTNT) from the L1 gene which would otherwise have caused transcription termination when expressed from a vaccinia virus early promoter. The nucleotide sequence was mutated to retain the correct amino acid sequence of the Ll protein. Full-length mRNA and Ll protein were generated in cells infected with the recombinant virus containing the mutant sequence, whereas the wild-type sequence generated only truncated mRNA and no detectable protein. Mice were immunized with VV expressing L1 from the mutant sequence and from the wild-type sequence in constructs with either early or late vaccinia virus promoters. Only the early promoter constructs were effective in priming cytotoxic T lymphocytes (CTL). Moreover the mutant sequence was significantly more effective than the wild-type sequence. The same L1 sequences, expressed from a vaccinia virus late promoter or coexpressed with MHC Class I molecules also expressed from a late promoter, produced high levels of Ll protein in both cases but nevertheless failed to elicit CTL activity. This is the first report of an HPV-specific CTL response and we have reaffirmed the importance of choosing the correct promoter and sequence expressed when using recombinant vaccinia viruses to induce cytotoxic T lymphocytes. These data are relevant for the design of vaccines to generate cell-mediated immunity against human papillomavirus infection.

Upstream promoter elements of the herpes simplex virus type 1 glycoprotein H gene

To investigate the cis-acting sequence elements that are involved in the regulation of herpes simplex virus type 1 late-gene expression, recombinant viruses were constructed that express the Escherichia coli lacZ gene from the promoter of the glycoprotein H (gH) gene. Deletion experiments established an upstream boundary for the gH promoter of no more than 83 bp from the start of gH transcription and showed that the promoter sequences did not overlap with coding sequences of the upstream thymidine kinase (tk) gene. Sequences of the tk gene previously shown to be required for efficient processing of the tk transcript were essential for expression form the gH promoter and included a TATA-like element. In addition, the gH TATA element was specifically mutagenized to substitute the TATA elements of immediate-early, early, and other late viral promoters for the gH TATA element. The results indicated that the TATA element was an interchangeable component of herpes simplex virus type 1 promoters and did not regulate temporal expression.

The target DNA sequence for resolution of poxvirus replicative intermediates is an active late promoter

The linear double-stranded genomes of poxviruses such as Shope fibroma virus (SFV) replicate autonomously within the cytoplasm of infected cells, and it is believed that all of the replication functions are virally encoded. During DNA replication the incompletely base-paired terminal hairpin loops of the viral genome transiently exist in the form of inverted repeat replicative intermediates. These inverted repeat structures form the target for telomere resolution events that include sequence-specific cleavage and directed strand exchange to form the hairpin termini of progeny virus genomes. The terminal sequence domain which forms the telomere resolution target (TRT) shares considerable sequence similarity with viral late promoters. In this study we demonstrate that the TRT of SFV is capable of functioning as a strong viral promoter late in infection. A spectrum of TRT mutations affects telomere resolution and late transcription in a strictly concordant fashion, suggesting that the two activities may be inextricably linked. Further support for this concept comes from the demonstration that a late SFV promoter sequence designated cryptic TRT, which differs substantially from the native TRT in terms of sequence, can support telomere resolution when placed in the correct spatial context. The proposed model for telomere resolution invokes directed unwinding of the TRT double helix by a transcription initiation complex and processing of the resulting secondary structure by viral late-gene products.

Purification of the DNA binding domain of herpes simplex virus type 1 immediate-early protein Vmw175 as a homodimer and extensive mutagenesis of its DNA recognition site.

The herpes simplex virus type 1 (HSV-1) Immediate-Early (IE) polypeptide Vmw175 is essential for the activation of transcription from viral early and late promoters. Vmw175 also reduces the activity of its own (IE-3) promoter in transfection assays. Both transactivation and repression mediated by Vmw175 require the integrity of a conserved domain of the polypeptide which has been shown to bind to specific DNA sequences. We have investigated the DNA sequence requirements for Vmw175 binding using a randomly mutated target. The results indicate that the binding site covers a region of 13 nucleotides divided into proximal and distal parts which are consistent with the consensus ATCGTNNNNNYSG. We have also expressed several different constructs encompassing the DNA binding domain of Vmw175 in bacteria, and obtained preparations of greater than 90% purity. The DNA binding domain is a dimer in solution, and binds DNA with a specificity similar to that of the intact protein, although the smallest DNA binding competent protein has a slightly reduced specificity.

Regulation of JCV promoter function: Evidence that a pentanucleotide “silencer” repeat sequence AGGGAAGGGA down-regulates transcription of the JC virus late promoter

The human neurotropic papovavirus JCV contains sequences within the two 98-bp tandem repeats which play a key role in glial-specific transcription of the viral early and late promoters. Previous analysis of the 98-bp sequence has delineated several protein-binding domains that are recognized by nuclear factors present in human brain cells. In the present study, by deletion mutation analysis, we have identified a region within each 98-bp repeat that reduces transcriptional activity of the JCV late promoter (JCV L). Using synthetic oligonucleotides spanning this region, designated “OP,” we demonstrate that down-regulation of the JCV L promoter is associated with a pentanucleotide repeat sequence (AGGGAAGGGA) juxtaposed to the poly(dA) tract within the 98-bp tandem repeats. The OP sequence interacts specifically with a protein derived from glial nuclear extract and forms a major 56- to 60-kDa complex. Methylation interference experiment indicates that the three G residues proximal to the poly(dA) tract make major groove contacts with the protein. Single-base-pair substitution of these residues suggests that the complex can form in the presence of two of the three guanosyl residues. The possible role of this protein in regulating the JCV lytic cycle in concert with nearby regulatory elements within JCV promoter region is discussed.

SV40 activates transcription from the transferrin receptor promoter by inducing a factor which binds to the CRE/AP-1 recognition sequence.

During the course of lytic infection by simian virus 40 (SV40), expression of both the viral late genes and certain host cellular genes is induced. The promoter of the cellular transferrin receptor (TR) gene contains a DNA sequence which is similar to the AP-1- and AP-4-binding region in SV40 which has been implicated in the control of the viral late promoter. Expression of TR is needed for cells to enter S-phase and is therefore expected to be important for the SV40 lytic cycle. Here we show that the level of TR mRNA in vivo was increased by SV40 infection. A factor which activates transcription from the TR promoter in vitro was specifically induced in SV40-infected cells. Gel mobility shift assays with an oligonucleotide comprising this part of the TR promoter showed three nucleoprotein complexes to be formed with proteins from CV-1 cells. Following SV40 infection, one of the complexes was increased ten-fold. Formation of this complex was specifically reduced by competition with the phorbol ester-responsive element of the collagenase gene, implying that the factor is a member of the AP-1/Jun/Fos family. Cross-linking of the complex by ultraviolet light showed major DNA-binding components to be proteins of about 55 kD and 47 kD. Removal of this factor by adding the oligonucleotide to in vitro transcription reactions with the TR promoter, abolished the activation of TR transcription. The factor which binds to the TR promoter co-sedimented with SV40 chromosomes extracted late in infection. This suggests that similar transcriptional regulatory proteins are involved in controlling transcription from both the SV40 and the TR promoters, and that the virus can use a common mechanism to induce viral and host cellular transcription.