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PMID: 2556265 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

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

The EMBO journal ·Vol. 8 ·No. 13 ·1989-12-20 ·Pages 4215-27

Macchi M, Bornert JM, Davidson I, Kanno M, Rosales R, Vigneron M, Xiao JH, Fromental C, Chambon P

Abstract

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.(ABSTRACT TRUNCATED AT 400 WORDS)

MeSH Terms
Animals Base Sequence Cell Line Cell Nucleus/metabolism Enhancer Elements, Genetic Genes, Viral H-2 Antigens/genetics HeLa Cells/metabolism Humans Molecular Sequence Data Mutation Nuclear Proteins/biosynthesis,metabolism Plasmids Promoter Regions, Genetic Protein Binding Restriction Mapping Simian virus 40/genetics
Chemicals
H-2 Antigens H-2Kb protein, mouse Nuclear Proteins
Authors & Affiliations
9 authors, click to expand affiliations / ORCID
Macchi M
Laboratoire de Génétique Moléculaire des Eucaryotes du CNRS, Unité 184 de Biologie Moléculaire et de Génie Génétique de l'INSERM, Institut de Chimie Biologique, Faculté de Médecine, Strasbourg, France.
Bornert J M
Davidson I
Kanno M
Rosales R
Vigneron M
Xiao J H
Fromental C
Chambon P
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1989-12-20
Pages
4215-27
Language
English
Region
England
NLM ID
8208664
PMCID
PMC401618
Subset
IM
External Links
PubMed source PMC Full Text View journal details
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