scholarly journals trans activation of the simian virus 40 late promoter by large T antigen requires binding sites for the cellular transcription factor TEF-1.

1991 ◽  
Vol 65 (12) ◽  
pp. 6535-6543 ◽  
Author(s):  
P Casaz ◽  
R Sundseth ◽  
U Hansen
1992 ◽  
Vol 12 (6) ◽  
pp. 2514-2524 ◽  
Author(s):  
Z S Guo ◽  
M L DePamphilis

The origins of DNA replication (ori) in simian virus 40 (SV40) and polyomavirus (Py) contain an auxiliary component (aux-2) composed of multiple transcription factor binding sites. To determine whether this component stimulated replication by binding specific transcription factors, aux-2 was replaced by synthetic oligonucleotides that bound a single transcription factor. Sp1 and T-antigen (T-ag) sites, which exist in the natural SV40 aux-2 sequence, provided approximately 75 and approximately 20%, respectively, of aux-2 activity when transfected into monkey cells. In cell extracts, only T-ag sites were active. AP1 binding sites could replace completely either SV40 or Py aux-2. Mutations that eliminated AP1 binding also eliminated AP1 stimulation of replication. Yeast GAL4 binding sites that strongly stimulated transcription in the presence of GAL4 proteins failed to stimulate SV40 DNA replication, although they did partially replace Py aux-2. Stimulation required the presence of proteins consisting of the GAL4 DNA binding domain fused to specific activation domains such as VP16 or c-Jun. These data demonstrate a clear role for transcription factors with specific activation domains in activating both SV40 and Py ori. However, no correlation was observed between the ability of specific proteins to stimulate promoter activity and their ability to stimulate origin activity. We propose that only transcription factors whose specific activation domains can interact with the T-ag initiation complex can stimulate SV40 and Py ori-core activity.


The transforming protein of Simian virus 40 (SV40), large T-antigen, regulates transcription both positively and negatively during the productive infection cycle. We have isolated a number of cellular genes which are expressed at elevated levels in SV40-transformed cells and have used these to study the mechanism or mechanisms by which the viral transforming protein regulates cellular gene expression. Small RNAs homologous to the mouse B2 repetitive sequence family are found at higher levels in transformed cells than in normal cells and we have shown that pure large T-antigen stimulates transcription of such repeats by RNA polymerase III. A class I major histocompatibility complex (MHC) gene is also activated as a result of SV40 transformation and we have used DNA-mediated gene transfer to study how this gene is regulated by large T-antigen.


1988 ◽  
Vol 8 (4) ◽  
pp. 1648-1656
Author(s):  
G J Gallo ◽  
G Gilinger ◽  
J C Alwine

The late promoter of simian virus 40 is transcriptionally activated, in trans, by large T antigen, the primary viral early gene product. Although large T antigen is a well-characterized DNA-binding protein, a variety of data suggest that its trans-activation function does not require direct interaction with DNA. We demonstrate that defined late promoter elements, omega (omega), tau (tau), and delta (delta), necessary for T-antigen-mediated trans-activation, are binding sites for simian cellular factors, not T antigen. Two of the late promoter elements (omega and tau) are shown to bind the same factor or family of factors. These factors bind to a site very similar to that for the HeLa cell factor AP1. We refer to these factors as the simian AP1-sequence recognition proteins (sAP1-SRPs). Compared with normal simian CV-1P cells, the sAP1-SRPs from T-antigen-producing COS cells, or from 14-h simian virus 40-infected CV-1P cells, showed altered binding patterns to both the omega and tau binding sites. In addition, the sAP1-SRPs from T-antigen-containing cells bound to the tau site more stably than did the analogous factors from normal CV-1P cells. The altered pattern of binding and the increased stability of binding correlated with the presence of T antigen in the cell. Additionally, the alteration of the binding pattern within 14 h of infection in CV-1P cells is temporally correct for late promoter activation. Overall, the data show (i) that the late promoter elements necessary for T-antigen-mediated trans-activation contain binding sites for simian cellular DNA-binding proteins; (ii) that the presence of T antigen causes alterations in the binding characteristics of specific simian cellular DNA-binding factors or families of factors; and (iii) that factors which bind to the late promoter elements required for activation have altered and more stable binding characteristics in the presence of T antigen. These points strongly suggest that T antigen mediates trans-activation indirectly through the alteration of binding of at least one specific simian cellular factor, sAP1-SRP, or through the induction of a family of sAP1-SRP factors.


1993 ◽  
Vol 13 (2) ◽  
pp. 961-969
Author(s):  
M C Gruda ◽  
J M Zabolotny ◽  
J H Xiao ◽  
I Davidson ◽  
J C Alwine

Simian virus 40 (SV40) large T antigen is a potent transcriptional activator of both viral and cellular promoters. Within the SV40 late promoter, a specific upstream element necessary for T-antigen transcriptional activation is the binding site for transcription-enhancing factor 1 (TEF-1). The promoter structure necessary for T-antigen-mediated transcriptional activation appears to be simple. For example, a promoter consisting of upstream TEF-1 binding sites (or other factor-binding sites) and a downstream TATA or initiator element is efficiently activated. It has been demonstrated that transcriptional activation by T antigen does not require direct binding to the DNA; thus, the most direct effect that T antigen could have on these simple promoters would be through protein-protein interactions with either upstream-bound transcription factors, the basal transcription complex, or both. To determine whether such interactions occur, full-length T antigen or segments of it was fused to the glutathione-binding site (GST fusions) or to the Gal4 DNA-binding domain (amino acids 1 to 147) (Gal4 fusions). With the GST fusions, it was found that TEF-1 and the TATA-binding protein (TBP) bound different regions of T antigen. A GST fusion containing amino acids 5 to 172 (region T1) efficiently bound TBP. TEF-1 bound neither region T1 nor a region between amino acids 168 and 373 (region T2); however, it bound efficiently to the combined region (T5) containing amino acids 5 to 383.(ABSTRACT TRUNCATED AT 250 WORDS)


1993 ◽  
Vol 13 (2) ◽  
pp. 961-969 ◽  
Author(s):  
M C Gruda ◽  
J M Zabolotny ◽  
J H Xiao ◽  
I Davidson ◽  
J C Alwine

Simian virus 40 (SV40) large T antigen is a potent transcriptional activator of both viral and cellular promoters. Within the SV40 late promoter, a specific upstream element necessary for T-antigen transcriptional activation is the binding site for transcription-enhancing factor 1 (TEF-1). The promoter structure necessary for T-antigen-mediated transcriptional activation appears to be simple. For example, a promoter consisting of upstream TEF-1 binding sites (or other factor-binding sites) and a downstream TATA or initiator element is efficiently activated. It has been demonstrated that transcriptional activation by T antigen does not require direct binding to the DNA; thus, the most direct effect that T antigen could have on these simple promoters would be through protein-protein interactions with either upstream-bound transcription factors, the basal transcription complex, or both. To determine whether such interactions occur, full-length T antigen or segments of it was fused to the glutathione-binding site (GST fusions) or to the Gal4 DNA-binding domain (amino acids 1 to 147) (Gal4 fusions). With the GST fusions, it was found that TEF-1 and the TATA-binding protein (TBP) bound different regions of T antigen. A GST fusion containing amino acids 5 to 172 (region T1) efficiently bound TBP. TEF-1 bound neither region T1 nor a region between amino acids 168 and 373 (region T2); however, it bound efficiently to the combined region (T5) containing amino acids 5 to 383.(ABSTRACT TRUNCATED AT 250 WORDS)


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