scholarly journals Transcription Factor Sp1 Mediates Cell-Specifictrans-Activation of the Human Cytomegalovirus DNA Polymerase Gene Promoter by Immediate-Early Protein IE86 in Glioblastoma U373MG Cells

1998 ◽  
Vol 72 (1) ◽  
pp. 236-244 ◽  
Author(s):  
Jun Wu ◽  
Joseph O’Neill ◽  
Miguel S. Barbosa
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Dna Polymerase ◽  
Human Cytomegalovirus ◽  
Binding Activity ◽  
Immediate Early ◽  
Polymerase Gene ◽  
Cellular Transcription Factor ◽  
Dna Binding Activity ◽  
Cellular Transcription

ABSTRACT Human cytomegalovirus (HCMV) gene expression is highly cell and tissue specific. Cell factor-mediated regulatory interactions are involved in regulating the restricted expression of the HCMV major immediate-early (IE) gene (J. F. Baskar, P. P. Smith, G. Nilaver, R. A. Jupp, S. Hoffmann, N. J. Peffer, D. J. Tenney, A. M. Colberg-Poley, P. Ghazal, and J. A. Nelson, 70:3207–3213, 1996). To gain an understanding of HCMV early gene activation, we studied the effect of each of the three major IE proteins, IE72, IE86, and IE55, on the HCMV DNA polymerase gene (pol; UL54) promoter. In transient-expression assays, the IE86 protein alone was able to transactivate the polpromoter, but IE72 and IE55 were not, in permissive U373MG cells. However, we were unable to detect IE86-mediated transactivation in nonpermissive HeLa or C33-A cells. Using electrophoretic mobility shift assays (EMSAs), we found that expression of the IE86 protein in U373MG cells resulted in specific binding of a DNA complex to an inverted-repeat element, IR1, of the pol promoter. Antibody supershifting and EMSA-Western blotting experiments further showed that IE86 and the cellular transcription factor Sp1 were components of the IR1 DNA-binding complex. Furthermore, we found that binding of DNA by Sp1 was dramatically increased in the presence of IE86. Interestingly, this IE86-induced DNA-binding activity of Sp1 was inhibited by a repressor activity presented in HeLa cells. In summary, our study suggests that a viral regulatory protein can modulate the DNA binding activity of a cellular transcription factor, resulting in cell-specific transactivation of viral genes.

scholarly journals Characteristics of DNA-Binding Activity of Human Cytomegalovirus ppUL44

2000 ◽  
Vol 44 (10) ◽  
pp. 827-832 ◽  
Author(s):  
Eung-soo Hwang ◽  
Jinhee Kim ◽  
Hyun-soon Jong ◽  
Jae-won Park ◽  
Chung-gyu Park ◽  
...  
Keyword(s):  
Dna Binding ◽  
Human Cytomegalovirus ◽  
Binding Activity ◽  
Dna Binding Activity

scholarly journals Suppression of E1A-Mediated Transformation by the p50E4F Transcription Factor

1999 ◽  
Vol 19 (7) ◽  
pp. 4739-4749 ◽  
Author(s):  
Elma R. Fernandes ◽  
Robert J. Rooney
Keyword(s):  
Cell Death ◽  
Transcription Factor ◽  
Suppressive Effect ◽  
Culture Conditions ◽  
Binding Activity ◽  
Cellular Transcription Factor ◽  
Dna Binding Activity ◽  
3T3 Fibroblasts ◽  
Induced Apoptosis ◽  
Nih 3T3

ABSTRACT The adenovirus E1A gene can act as an oncogene or a tumor suppressor, with the latter effect generally arising from the induction of apoptosis or the repression of genes that provide oncogenic growth stimuli (e.g., HER-2/c-erbB2/neu) or increased metastatic invasiveness (e.g., metalloproteases). In this study, coexpression of E1A and p50E4F, a cellular transcription factor whose DNA binding activity is stimulated by E1A, suppressed colony formation by NIH 3T3 cells and transformation of primary rat embryo fibroblasts but had no observed effect in the absence of E1A. Domains in p50E4F required for stimulation of the adenovirus E4 promoter were required for the suppressive effect, indicating a transcriptional mechanism. In serum-containing media, retroviral expression of p50E4F in E1A13S/ras-transformed NIH 3T3 fibroblasts had little effect on subconfluent cultures but accelerated a decline in viability after the cultures reached confluence. Cell death occurred by both apoptosis and necrosis, with the predominance of each process determined by culture conditions. In serum-free media, p50E4F accelerated E1A-induced apoptosis. The results suggest that p50E4F sensitizes cells to signals or conditions that cause cell death.

A single polypeptide possesses the binding and transcription activities of the adenovirus major late transcription factor

1986 ◽  
Vol 6 (12) ◽  
pp. 4723-4733
Author(s):  
L A Chodosh ◽  
R W Carthew ◽  
P A Sharp
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Binding Site ◽  
Rna Polymerase Ii ◽  
Sodium Dodecyl ◽  
Binding Activity ◽  
Affinity Column ◽  
Dna Binding Activity ◽  
Dna Fragment

A simple approach has been developed for the unambiguous identification and purification of sequence-specific DNA-binding proteins solely on the basis of their ability to bind selectively to their target sequences. Four independent methods were used to identify the promoter-specific RNA polymerase II transcription factor MLTF as a 46-kilodalton (kDa) polypeptide. First, a 46-kDa protein was specifically cross-linked by UV irradiation to a body-labeled DNA fragment containing the MLTF binding site. Second, MLTF sedimented through glycerol gradients at a rate corresponding to a protein of native molecular weight 45,000 to 50,000. Third, a 46-kDa protein was specifically retained on a biotin-streptavidin matrix only when the DNA fragment coupled to the matrix contained the MLTF binding site. Finally, proteins from the most highly purified fraction which were eluted and renatured from the 44- to 48-kDa region of a sodium dodecyl sulfate-polyacrylamide gel exhibited both binding and transcription-stimulatory activities. The DNA-binding activity was purified 100,000-fold by chromatography through three conventional columns plus a DNA affinity column. Purified MLTF was characterized with respect to the kinetic and thermodynamic properties of DNA binding. These parameters indicate a high degree of occupancy of MLTF binding sites in vivo.

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