The SV40 TC‐II(kappa B) enhanson binds ubiquitous and cell type specifically inducible nuclear proteins from lymphoid and non‐lymphoid cell lines

We have characterized the complexes resulting from the specific binding in vitro of proteins present in nuclear extracts of several lymphoid and non‐lymphoid cell lines to the TC‐I and TC‐II sequences of the simian virus 40 (SV40) enhancer. No proteins could be detected, binding selectively to the T...

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Bibliographic Details
Published in:The EMBO journal 1989-12, Vol.8 (13), p.4215-4227
Main Authors: Macchi, M., Bornert, J.M., Davidson, I., Kanno, M., Rosales, R., Vigneron, M., Xiao, J.H., Fromental, C., Chambon, P.
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Biological and medical sciences
Cell Line
Cell Nucleus - metabolism
Enhancer Elements, Genetic
Fundamental and applied biological sciences. Psychology
Genes, Viral
H-2 Antigens - genetics
HeLa Cells - metabolism
Humans
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
Mutation
Nuclear Proteins - biosynthesis
Nuclear Proteins - metabolism
Plasmids
Promoter Regions, Genetic
Protein Binding
Restriction Mapping
Simian virus 40 - genetics
Transcription. Transcription factor. Splicing. Rna processing
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Summary:We have characterized the complexes resulting from the specific binding in vitro of proteins present in nuclear extracts of several lymphoid and non‐lymphoid cell lines to the TC‐I and TC‐II sequences of the simian virus 40 (SV40) enhancer. No proteins could be detected, binding selectively to the TC‐I sequence, but two proteins TC‐IIA and TC‐IIB were identified interacting specifically with both the TC‐II/kappa B enhanson, 5′‐GGAAAGTCCCC‐3′ (important for the activity of the SV40 enhancer in vivo), and with the related H‐2Kb enhanson, 5′‐TGGGGATTCCCCA‐3′. The binding of these two proteins to mutated TC‐II enhansons correlates with the effect of these mutations in vivo, suggesting that both proteins may be important for SV40 enhancer activity. The TC‐IIA binding activity was present in nuclear extracts of mature lymphoid B cells and was increased in pre‐B cell nuclear extracts by lipopolysaccharide (LPS) and cycloheximide treatment. Furthermore, complex formation between the TC‐IIA protein and the TC‐II enhanson was efficiently competed by the kappa B motif from the kappa chain enhancer, indicating that TC‐IIA is the NF‐kappa B factor or a closely related protein. However, in contrast to previous reports, a TC‐IIA/NF‐kappa B‐like protein whose properties could not be distinguished from those of the TC‐IIA protein present in lymphoid B cells, was found in nuclear extracts of several untreated non‐lymphoid cell lines, notably of HeLa cells, but not of undifferentiated F9 embryonal carcinoma (EC) cells [F9(ND)]. The TC‐IIA binding activity which was moderately increased in HeLa cell nuclear extracts by 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA) and/or cycloheximide treatment could be induced in nuclear extracts of F9(ND) cells by cycloheximide, but not by TPA. Moreover, the TC‐IIA binding activity could be induced in cytosolic fractions from F9(ND) cells by treatment with deoxycholate, indicating that these cells contain an inhibitor protein similar to the previously described NF‐kappa B inhibitor, I kappa B. The second TC‐II enhanson binding protein, TC‐IIB, which could be clearly distinguished from the TC‐IIA/NF‐kappa B‐like protein, by a number of differential properties, resembles the previously described KBF1/H2TF1 protein as it binds with a higher affinity to the H‐2Kb enhanson than to the TC‐II/kappa B enhanson, and its pattern of methylation interference on the H‐2Kb and TC‐II/kappa B enhansons is identical to that reported for the KBF1/H2TF1 protein.
ISSN:0261-4189
1460-2075
DOI:10.1002/j.1460-2075.1989.tb08607.x