Octamer Binding Site Research Articles

Binding and cooperative interactions between two B cell-specific transcriptional coactivators.

The class II transactivator (CIITA) and B cell octamer-binding protein 1/octamer-binding factor 1/Oct coactivator from B cells (Bob1/OBF-1/OCA-B) represent two B cell-specific transcriptional coactivators. CIITA and Bob1 interact with proteins that bind to conserved upstream sequences in promoters of class II major histocompatibility genes and octamer-binding transcription factors Oct-1 and Oct-2, respectively. Both CIITA and Bob1 increase the expression from the DRA promoter, which is a prototypic class II promoter. Moreover, in the presence of CIITA, interactions between class II promoters and Bob1 are independent of the octamer-binding site. Using in vivo and in vitro binding assays, we confirm that Bob1 binds to CIITA. Thus, CIITA not only activates the expression of class II genes but recruits another B cell-specific coactivator to increase transcriptional activity of class II promoters in B cells.

Oct-1 [corrected] and Oct-2 DNA-binding site specificity is regulated in vitro by different kinases.

The transcription factors Oct-1 and Oct-2 bind differentially to three octamer binding sequences corresponding to the octamer binding site from the H2B promoter [ATGCTAATAA], a simple TAATGARAT motif, found in herpes simplex virus IE4/5 genes [GCGGTAATGAGAT], and a perfect consensus overlapping octamer/TAATGARAT motif [ATGCTAATGAGAT]. By comparing the effects of protein kinase A, protein kinase C and casein kinase 2 in vitro on the binding of Oct-1 and Oct-2 to the three motifs, we show that the actions of these kinases regulate Oct-1 and Oct-2 DNA binding independently of each other in a binding-site-specific manner. Inhibition of cellular phosphatases also regulate Oct-1 and Oct-2 DNA binding in a binding-site-specific manner. Both kinase and phosphatase activity are important for regulating the DNA binding activity of Oct-1 and Oct-2 because, in the presence of phosphatase inhibitors, protein kinase A attenuates the binding of both Oct-1 and Oct-2 to the octamer binding site but enhances binding when phosphatase inhibitors are omitted. Thus the DNA specificity of Oct-1 and Oct-2 can be regulated in vitro by the action of different kinases.

Interaction of Oct-1 with TFIIB: IMPLICATIONS FOR A NOVEL RESPONSE ELICITED THROUGH THE PROXIMAL OCTAMER SITE OF THE LIPOPROTEIN LIPASE PROMOTER

The ubiquitous human POU domain protein, Oct-1, and the related B-cell protein, Oct-2, regulate transcription from a variety of eukaryotic genes by binding to a common cis-acting octamer element, 5'-ATTTGCAT-3'. The binding of Oct-1 and Oct-2 to the functionally important lipoprotein lipase (LPL) promoter octamer site was stimulated by the general transcription factor, TFIIB. Comparative analysis of the LPL, histone H2B (H2B), and herpes simplex virus ICPO gene promoter octamer sites revealed that nucleotide sequences within and flanking the octamer sequence determined the degree of TFIIB-mediated stimulation of Oct-1 DNA binding. TFIIB was found to decrease the rate of dissociation of Oct-1 from the LPL octamer site, whereas it increased the rate of association, as well as decreased the rate of dissociation, of Oct-1 from the H2B octamer site. A monoclonal antibody against TFIIB immunoprecipitated a ternary complex containing TFIIB, Oct-1, and the LPL and H2B octamer binding sites. TFIIB did not alter the DNase I footprints generated by Oct-1 on the LPL and H2B promoters. However, Oct-1 on the TATA-binding protein and TFIIB from footprinting the perfect TATA box sequence located 5' of the LPL, NF-Y binding site. In transfection experiments, transcription from the reporters containing the LPL octamer, and either the SV40 or the yeast transcription factor GAL4-dependent enhancers, initiated at a precise position within the octamer sequence. Transcription from reporters containing the H2B octamer and the SV40 enhancer initiated at several positions within and flanking the octamer site, whereas transcription initiated at a precise position within the octamer from reporters with both the H2B octamer and the GAL4-dependent enhancer. These results suggest that octamers and their flanking sequences play an important role in positioning the site of transcription initiation, and that this could be a function of the interaction of Oct-1 with TFIIB.

Regulatory function of the equine herpesvirus 1 ICP27 gene product

The UL3 protein of equine herpesvirus 1 (EHV-1) KyA strain is a homolog of the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1) and the ORF 4 protein of varicella-zoster virus. To characterize the regulatory function of the UL3 gene product, a UL3 gene expression vector (pSVUL3) and a vector expressing a truncated version of the UL3 gene (pSVUL3P) were generated. These effector plasmids, in combination with an EHV-1 immediate-early (IE) gene expression vector (pSVIE) and chimeric EHV-1 promoter-chloramphenicol acetyltransferase (CAT) reporter constructs, were used in transient transfection assays. These assays demonstrated that the EHV-1 UL3 gene product is a regulatory protein that can independently trans activate the EHV-1 IE promoter; however, this effect can be inhibited by the repressive function of the IE gene product on the IE promoter (R. H. Smith, G. B. Caughman, and D. J. O'Callaghan, J. Virol. 66:936-945, 1992). In the presence of the IE gene product, the UL3 gene product significantly augments gene expression directed by the promoters of three EHV-1 early genes (thymidine kinase; IR4, which is the homolog of HSV-1 ICP22; and UL3 [ICP27]) and the promoter of the EHV-1 late gene IR5, which is the homolog of HSV-1 US10. Sequences located at nucleotides -123 to +20 of the UL3 promoter harbor a TATA box, SP1 binding site, CAAT box, and octamer binding site and, when linked to the CAT reporter gene, are trans activated to maximal levels by the pSVIE construct in transient expression assays. Results from CAT assays also suggest that the first 11 amino acids of the UL3 protein are not essential for the regulatory function of the UL3 gene product.

Characterization of the human interleukin‐5 gene promoter: involvement of octamer binding sites in the gene promoter activity

To investigate the human interleukin (IL)-5 gene promoter, we have constructed a plasmid with the firefly luciferase reporter gene linked to human IL-5 5' flanking sequence (nucleotides -507 to +44). We have used this plasmid to transfect the mouse EL4 T cell line, which can, under certain conditions, produce IL-5 transcripts. Phorbol 12-myristate 13-acetate, A23187 and N6, 2'-O-dibutyryl-adenosine 3':5'-cyclic monophosphate co-stimulation of EL4 cells transfected with the human IL-5/luciferase reporter gene construct resulted in maximal induction of the luciferase gene. Deletion analysis of the IL-5 promoter revealed the presence of negative regulatory elements between nucleotides -404 and -312 and two regions, located between nucleotides -312 and -227 and between nucleotides -80 and -35, that are involved in the positive regulation of the IL-5 promoter. Using electrophoretic mobility shift assays, we show that the positive element located between nucleotides -312 and -227 involves the binding of factors antigenically related to Oct1, Oct2A and Oct2B, to a perfect octamer motif located at position -244/-237. Introduction of three point mutations in the octamer motif of the IL-5/luciferase reporter gene plasmid, which results in the loss of competition for the factors binding to the IL-5 promoter sequence, reduced the production of luciferase from stimulated, transfected EL4 cells, by 90%. Octamer factors can also bind within the second positive regulatory region.

Octamer factors exert a dual effect on the IL-2 and IL-4 promoters.

The promoters of IL-2 and IL-4 genes contain multiple binding sites for octamer factors. In peripheral T lymphocytes and several T cell lines, both the ubiquitous Oct factor Oct-1 and the lymphocyte-specific factor Oct-2 are expressed and bind to the IL-2 and IL-4 promoters. Prominent octamer binding sites of IL-2 and IL-4 promoters are their upstream promoter sites (UPS) which share 14 identical nucleotides. Multiple copies of the IL-2 and IL-4 UPS act as inducible enhancers in T cells, and their induction is inhibited by the immunosuppressant cyclosporin A (CsA). Closely linked to the octamer site, the IL-2 UPS contains a non-canonical AP-1 binding (TRE) site, and mutation in either site to a non-functional factor binding site impairs the induction of the IL-2 promoter. The binding of AP-1 and octamer factors to the IL-2 UPS DNA overlaps, and the tight association and functional cooperation of octamer with AP-1 factors is of crucial importance for the inducible IL-2 UPS activity. Introduction of five or ten spacer nucleotides between both IL-2 UPS sites results in a drastic reduction of inducible UPS activity, both in the loss of suppression by CsA and stimulation by the Ca(2+)-dependent phosphatase calcineurin. Within the IL-4 UPS the Oct and TRE-like motifs are separated by a binding site of nuclear factor of activated T cells (NF-AT). This site shares nine out of ten bp with an IL-2 NF-AT site. The strong binding of NF-ATp to the IL-4 UPS site suppresses the simultaneous binding of Oct factors to the IL-4 UPS. Because the two other Oct binding sites of IL-4 promoter show a similar sequence configuration, the binding of NF-AT seems to prevent the simultaneous binding of Oct factors to the IL-4 promoter. By contrast, both classes of factors bind simultaneously to the IL-2 promoter, and their tight association with AP-1 enhances the IL-2 promoter activity.

Cloning of the Xenopus laevis cdk2 promoter and functional analysis in oocytes and during early development

CDK2 (cyclin-dependent kinase 2) is a serine/threonine kinase which is involved in regulating S-phase entry in higher eukaryotes. To investigate the transcriptional control of this gene, a 13-kb Xenopus laevis genomic clone containing the 5' flanking sequences was isolated. A 2.7-kb fragment containing the promoter region was sequenced and the transcription start point (tsp) was determined by primer extension. Several putative regulatory elements, such as the E2F-binding site, Y box and octamer-binding site, were localized in this region, but no TATA box was found. When fused to cat, a reporter gene encoding chloramphenicol acetyltransferase, the 5' flanking sequences were shown to function in oocytes and an enhancer activity was found in this region. During early embryogenesis, cdk2 promoter activity was tested and de novo transcription was detected at the mid-blastula transition.

Positive and negative regulation of the composite octamer motif of the interleukin 2 enhancer by AP-1, Oct-2, and retinoic acid receptor.

The differentiating agent retinoic acid (RA) has been previously reported to interfere with 12-O-tetradecanoyl-phorbol-13-acetate (TPA)/Ca2+-induced signals for the regulation of the -96 to -66-bp octamer motif found in the enhancer for the interleukin (IL)-2 gene, which encodes a major T lymphocyte growth factor. The IL-2 octamer motif is a composite cis-element which binds Oct-1 and Oct-2 as well as a TPA/Ca2+-inducible nuclear factor, previously termed octamer-associated protein (OAP40). We show here that Oct-2, despite the presence of an active transcriptional activation domain, requires TPA/Ca2+-induced signals to strongly transactivate the IL-2 octamer motif in Jurkat T cells. This Oct-2-dependent transactivation is inhibited by RA. The presence of an intact COOH-terminal domain of Oct-2 contributes to both TPA/Ca2+-induced transactivation and the RA-mediated repression. We also show that both Fos and Jun components of the AP-1 factors participate in the OAP40 complex. Furthermore, transfected c-jun, jun-B, jun-D, c-fos, or Fos-B expression vectors partially substitute for TPA and Ca2+ and cooperate with Oct-2 for the transactivation of the combined OAP/octamer cis-element. Mutations of the genuine octamer-binding site abrogate both the binding of Oct-1 and Oct-2 and the TPA/Ca2+-induced transactivation of the OAP/octamer motif. OAP confers to Oct-2 responsivity to both TPA/Ca2+ and RA, since specific mutations of the AP-1/OAP-binding site significantly reduce the transactivation by Oct-2 in response to TPA and Ca2+ and abolish the inhibition by RA. Furthermore, retinoic acid receptor (RAR) alpha is able to inhibit in vitro the formation of the complex between the nuclear AP-1/OAP and its specific binding site, resulting in the interference with Oct-2-dependent cis-regulatory function of this AP-1 element. Therefore, we propose that the TPA/calcium-activated AP-1/OAP element is the main target of positive or negative regulatory signals influencing the IL-2 octamer motif, through synergism with Oct-2 and antagonism by RAR.

Nucleosome Positioning on Chicken and Human Globin Gene Promoters in Vitro: Novel Mapping Techniques

We have developed two new techniques to assess the positions adopted by core histone octamers when reconstituted onto DNA. These, together with a previously described technique, were applied to mapping binding sites on plasmid DNAs containing either the human ζ-globin or chicken β-globin gene promoters. Each of the approaches enabled the sites occupied by histone octamers to be measured at high resolution and, in qualitative terms, revealed the same pattern of multiple, overlapping sites. Monomer extension, one of the novel techniques, can be used to reveal binding sites over extensive stretches of a single reconstitute (≈ 1000 bp). We found the distribution of histone octamer binding sites to be largely independent of the conditions employed for reconstitution, the topology of the DNA substrate and prolonged incubation under various post-reconstitution conditions. These properties, and features of the binding site maps that we derived, suggest that histone octamer positioning on these DNAs is predominantly a characteristic of the DNA sequence itself and, by implication, that nucleosome—nucleosome interactions and the formation of nucleosome arrays are of minor influence. Some of the techniques provide quantitative information concerning the relative binding strengths of the core histone octamer for different positioning sequences. In this context, it is notable that the majority of potential binding sites compete very poorly for the histone octamer, demonstrating that under the conditions pertinent to our analysis, the range of binding strengths exhibited by the octamer for particular DNA sequences is extensive, and greater than that observed when competitive binding has been studied by methods that do not reflect precise positioning.

Endothelial-cell-specific regulation of von Willebrand factor gene expression.

In both tissue sections and cell culture, the endothelial nature of a cell is most commonly determined by demonstration of its expression of von Willebrand factor (vWf) protein and/or mRNA. Thus, the mechanism of cell-type-specific transcriptional regulation of the vWf gene is central to studying the basis of endothelial-cell-specific gene expression. In this study, deletion analyses were carried out to identify the region of the vWf gene which regulates its endothelial-cell-specific expression. A 734-bp fragment which spans the sequence from -487 to +247 relative to the transcription start site was identified as the cell-type-specific promoter. It consists of a minimal core promoter located between -90 and +22, a strong negative regulatory element located upstream of the core promoter (ca. -500 to -300), and a positive regulatory region located downstream of the core promoter in the first exon. The activity of the core promoter is not cell type specific, and the negative regulatory region is required to inhibit its activity in all cell types. The positive regulatory region relieves this inhibition only in endothelial cells and results in endothelial-cell-specific gene expression. The positive regulatory region contains sequences predicting possible SP1, GATA, and octamer binding sites. Mutations in either the SP1 or octamer sequence have no effect on transcriptional activity, while mutation in the GATA binding element totally abolishes the promoter activity. Evidence that a GATA factor is involved in this interaction is presented. Thus, the positive regulatory region with an intact GATA binding site is required to overcome the inhibitory effect of the negative regulatory element and activate vWf gene expression in an endothelial-cell-specific manner.

Endothelial-cell-specific regulation of von Willebrand factor gene expression.

In both tissue sections and cell culture, the endothelial nature of a cell is most commonly determined by demonstration of its expression of von Willebrand factor (vWf) protein and/or mRNA. Thus, the mechanism of cell-type-specific transcriptional regulation of the vWf gene is central to studying the basis of endothelial-cell-specific gene expression. In this study, deletion analyses were carried out to identify the region of the vWf gene which regulates its endothelial-cell-specific expression. A 734-bp fragment which spans the sequence from -487 to +247 relative to the transcription start site was identified as the cell-type-specific promoter. It consists of a minimal core promoter located between -90 and +22, a strong negative regulatory element located upstream of the core promoter (ca. -500 to -300), and a positive regulatory region located downstream of the core promoter in the first exon. The activity of the core promoter is not cell type specific, and the negative regulatory region is required to inhibit its activity in all cell types. The positive regulatory region relieves this inhibition only in endothelial cells and results in endothelial-cell-specific gene expression. The positive regulatory region contains sequences predicting possible SP1, GATA, and octamer binding sites. Mutations in either the SP1 or octamer sequence have no effect on transcriptional activity, while mutation in the GATA binding element totally abolishes the promoter activity. Evidence that a GATA factor is involved in this interaction is presented. Thus, the positive regulatory region with an intact GATA binding site is required to overcome the inhibitory effect of the negative regulatory element and activate vWf gene expression in an endothelial-cell-specific manner.

Vascular cell adhesion molecule 1: contrasting transcriptional control mechanisms in muscle and endothelium.

Interaction between vascular cell adhesion molecule 1 (VCAM-1), which appears on the surface of endothelial cells in response to inflammation, and its integrin counter receptor, alpha 4 beta 1, on immune cells is responsible for targeting these immune cells to cytokine-stimulated endothelium. In addition to its role in the immune system, VCAM-1 is also expressed in a developmentally specific pattern on differentiating skeletal muscle, where it mediates cell-cell interactions important for myogenesis through interaction with alpha 4 beta 1. In contrast to endothelium, there is high basal expression of VCAM-1 in skeletal muscle cells and the expression is not cytokine-responsive. Here, we examine the molecular basis for these contrasting patterns of expression in muscle and endothelium, using VCAM-1 promoter constructs in a series of transfection assays. In endothelial cells, octamer binding sites act as silencers that prevent VCAM-1 expression in unstimulated cells. Tumor necrosis factor alpha overcomes the negative effects of these octamers and activates the promoter through two adjacent NF-kappa B binding sites. In muscle cells, a position-specific enhancer located between bp -21 and -5 overrides the effect of other promoter elements, resulting in constitutive VCAM-1 expression. A nuclear protein binds the position-specific enhancer in muscle but not endothelial cells; thus the pattern of expression of this protein could control enhancer activity.

Transforming growth factor beta and cyclosporin A inhibit the inducible activity of the interleukin-2 gene in T cells through a noncanonical octamer-binding site.

Transforming growth factor beta (TGF-beta) has a growth-inhibitory effect on numerous different cell types of the immune system, including T lymphocytes. We show in this study that the inhibitory action of TGF-beta on T lymphocytes is accompanied by a block of interleukin 2 (IL-2) gene expression which is mediated, at least in part, by inhibition of IL-2 promoter/enhancer activity. The functional analysis of cis-regulatory (proto-enhancer) elements of the IL-2 enhancer/promoter region showed that the most TGF-beta-responsive element maps to its so-called upstream promoter site. The proto-enhancer activity of the upstream promoter site element is also inhibited by cyclosporin A. The upstream promoter site DNA harbors two noncanonical, closely linked binding sequences for octamer and AP-1-like factors. Both sites are involved in the establishment of IL-2 enhancer activity. Since the activity of genuine octamer sites but not that of AP-1-binding sites is also impaired by TGF-beta and cyclosporin A in El4 T lymphoma cells, we conclude that both immunosuppressives interfere with the activity but not the DNA binding of octamer factors in T lymphocytes.

Transforming growth factor beta and cyclosporin A inhibit the inducible activity of the interleukin-2 gene in T cells through a noncanonical octamer-binding site.

Transforming growth factor beta (TGF-beta) has a growth-inhibitory effect on numerous different cell types of the immune system, including T lymphocytes. We show in this study that the inhibitory action of TGF-beta on T lymphocytes is accompanied by a block of interleukin 2 (IL-2) gene expression which is mediated, at least in part, by inhibition of IL-2 promoter/enhancer activity. The functional analysis of cis-regulatory (proto-enhancer) elements of the IL-2 enhancer/promoter region showed that the most TGF-beta-responsive element maps to its so-called upstream promoter site. The proto-enhancer activity of the upstream promoter site element is also inhibited by cyclosporin A. The upstream promoter site DNA harbors two noncanonical, closely linked binding sequences for octamer and AP-1-like factors. Both sites are involved in the establishment of IL-2 enhancer activity. Since the activity of genuine octamer sites but not that of AP-1-binding sites is also impaired by TGF-beta and cyclosporin A in El4 T lymphoma cells, we conclude that both immunosuppressives interfere with the activity but not the DNA binding of octamer factors in T lymphocytes.

The octamer binding site in the HPV16 regulatory region produces opposite effects on gene expression in cervical and non-cervical cells.

The upstream regulatory region (URR) of the tumorigenic human papillomaviruses HPV 16 and 18 contains an octamer binding site which is located adjacent to a binding site for the ubiquitous transcription factor NFI. The octamer site binds both the constitutively expressed transcription factor Oct-1 and a novel cervical octamer binding protein. In contrast the URR of the non-tumorigenic viruses HPV6 and HPV11 lacks the octamer binding site although the adjacent NFI site is conserved. Inactivation of the octamer binding site results in a higher level of gene expression in cells which contain only Oct-1 and a lower level in cells containing the cervical octamer binding protein indicating that that whilst Oct-1 binding reduces promoter activity, the cervical protein increases it. In agreement with this, over-expression of Oct-1 reduces the level of gene activity directed by this region of the HPV 16/18 URR and inhibits its activation by NFI whilst having no effect on the corresponding region of the HPV 6/11 URR. The significance of these effects is discussed in terms of the cervical-specific activity of the HPV16/18 URR and its role in HPV-mediated transformation.

Multiple octamer binding sites in the promoter region of the bovine alpha s2-casein gene.

Using a set of overlapping oligonucleotides from the promoter region of the bovine alpha s2-casein gene we have identified two nuclear factors which probably are involved in expression of this gene and the related calcium sensitive alpha s1- and beta-casein genes. One of these factors which was present in extracts of all tissues that have been tested including Hela cells turned out to be the octamer binding protein OCT-1. Oct-1 binds with different affinity to 4 sites at positions centred around -480, -260, -210 and -50. The strongest of these 4 binding sites, the one around position -50, is highly conserved in all calcium sensitive caseins of mouse, rat, rabbit and cattle. The other nuclear factor (MGF, mammary gland factor) which is specifically expressed in the mammary gland, binds to a site around position -90. This binding site is also highly conserved in all calcium sensitive caseins of mouse, rat, rabbit and cattle.

23] Algorithms for prediction of histone octamer binding sites

Publisher Summary This chapter presents algorithms for the prediction of histone octamer binding sites. Methods for the prediction of preferred binding sites for the histone octamer on a defined DNA sequence are of two types. In statistically based algorithms, the tested sequences are compared against the sequence organization of experimentally determined binding sites. By contrast, for structurally based algorithms, the preferred configuration of a sequence is calculated based on known and inferred structures of individual base steps. This approach is essentially a calculation of preferred curvature. The direction of the local axis of the double helix will change whenever the planes of adjacent base pairs are inclined relative to each other. The assignment of this principal component of curvature—the “wedge” angle of a base step—depends heavily on the limited number of available crystal structures of short DNA oligomers. This wedge angle has two components, a relative roll and a relative tilt. Of these two components, the relative roll is generally substantially larger and energetically more favorable. Therefore, in some algorithms relative tilt is justifiably ignored. Where these parameters are not available from crystal structures, they are estimated either from conformational energy calculations or by reference to rotational sequence preferences. An alternative and less direct approach is to calculate the wedge angles from the anomalous gel mobilities of a variety of DNA fragments of different sequence. These methods yield widely disparate values for particular individual base steps.

Transcription factor requirements for U2 snRNA-encoding gene activation in B lymphoid cells

Transcription of a human U2 small nuclear RNA(snRNA)-encoding gene in HeLa cells requires a distal enhancer element, which is composed of one octamer motif ( Oct) and three Sp1-binding sites. To study the transcription factor requirement in B-cells, different U2 enhancer constructions were transfected into the lymphoid cell line, BJA-B. The results showed that the activation of U2 snRNA transcription in B-cells also requires an enhancer comprising both the Oct and at least one Sp1-binding site. Deletion of all the Sp1-binding sites from the enhancer reduces transcription by 80–90% in HeLa, as well as in BJA-B cells, whereas the removal of the octamer-binding site reduces transcription to levels below detection in both cell types. Enhancers containing a single Oct have, nevertheless, the capacity to partially activate U2 snRNA transcription in both HeLa cells, in which only OTF-1 is expressed, and in BJA-B cells in which OTF-2 is the predominantly expressed octamer-binding factor. The most likely interpretation of our results is that both the ubiquitous transcription factor, OTF-1, and the B-cell-specific transcription factor, OTF-2, can activate U2 snRNA transcription. The results also revealed a similar functional cooperation between the transcription factors which bind to the Oct and the adjacent Sp1-binding site in BJA-B cells, as has been observed in HeLa cells, since a template which contains a weak binding site for OTFs expresses wild-type levels of U2 snRNA in both cell types when the weak octamer-binding site is combined with a Sp1-binding site.

Both Oct-1 and Oct-2A contain domains which can activate the ubiquitously expressed U2 snRNA genes.

The U2 snRNA genes, which are transcribed by RNA polymerase II at high levels in all tissues examined, require both a distal and a proximal sequence element for efficient expression. The distal sequence element which has many properties in common with transcriptional enhancers contains, in addition to Sp1 binding sites, an octamer binding site which mediates activation through interactions with the ubiquitous transcription factor Oct-1. In the present study we have attempted to answer the question whether Oct-1 contains a unique activating domain which is required for activation of snRNA genes or whether ubiquitously expressed and lymphoid specific octamer binding factors both have the capacity to activate snRNA transcription. Our results show that in the presence of Oct-1, overexpression of Oct-2A in HeLa or COS1 cells neither inhibits nor stimulates transcription of U2 constructions which contain octamer binding sites with or without an adjacent Sp1 binding site. Moreover, an Oct-2A--GAL4 fusion protein in which the DNA binding domain of Oct-2A was substituted for by the one of the yeast transcription activator GAL4 activates transcription of a human U2 snRNA gene in which the octamer binding site was replaced by a GAL4 binding site. From the results it is concluded that both Oct-1 and Oct-2A contain domains which can activate the ubiquitously expressed U2 snRNA genes.

The herpes simplex virus virion protein Vmw65 transcriptionally activates the gene encoding the U4 snRNA but not that encoding the U2 snRNA during lytic infection

Although lytic infection with herpes simplex virus (HSV) causes the repression of most host cell biosynthesis, it results in increased transcription of the cellular gene encoding the U4 snRNA, leading to accumulation of this snRNA. In contrast, no increased transcription of the gene encoding the U2 snRNA or accumulation of this RNA is observed in infected cells. These effects are mediated by the HSV virion protein Vmw65, which activates the U4 gene but does not affect the U2 gene. The significance of this difference between the U2 and U4 genes is discussed with regard to the presence in both of these genes of an identical octamer-binding site for the cellular transcription factor Oct-1 which complexes with Vmw65.