A nonerythroid GATA-binding protein is required for function of the human preproendothelin-1 promoter in endothelial cells

1990 ◽  
Vol 10 (9) ◽  
pp. 4854-4862
Author(s):  
D B Wilson ◽  
D M Dorfman ◽  
S H Orkin
Keyword(s):  
Growth Hormone ◽  
Endothelial Cells ◽  
Amino Acid ◽  
Dna Binding ◽  
Binding Protein ◽  
Cell Types ◽  
Dna Binding Protein ◽  
Core Motif ◽  
Transcription Activity ◽  
Shift Analysis

Endothelin-1 (ET-1) is a 21-amino-acid peptide synthesized by endothelial cells that has potent vasoconstrictor activity. Human ET-1 is derived from a 212-amino-acid prepropeptide, termed preproendothelin-1 (PPET-1). To identify cis-acting sequences essential for PPET-1 gene transcription, bovine aortic endothelial (BAE) cells were transfected with plasmids containing 5'-flanking sequences of the human PPET-1 gene fused to the human growth hormone gene as a reporter. Deletional analysis of these fusion plasmids showed that the sequence spanning positions -141 to -127 of the human PPET-1 promoter is required for full transcription activity. Introduction of clustered point mutations into this region of the promoter reduced transcription activity. Gel shift analysis, methylation interference, protein-DNA cross-linking, and oligonucleotide competition studies revealed that BAE cell nuclear extract contains a 47-kilodalton DNA-binding protein recognizing the core motif TATC (GATA) located at positions -135 to -132 of the PPET-1 promoter. The size and specificity of this DNA-binding protein resemble GF-1, a previously described transcription factor of erythroid cells that binds to the same core motif. Gel shift analysis indicated that GF-1 and the DNA-binding protein interacting with the PPET-1 promoter have different tissue distributions; the former is restricted to a subset of hematopoietic cells, and the latter is found in various cell types, including BAE, NIH 3T3, and HeLa cells. By using an antiserum to the C-terminal region of GF-1, the two proteins were also found to be antigenically distinct. When a growth hormone fusion plasmid containing the proximal 141 nucleotides of the PPET-1 promoter was transfected into a variety of cell types, these was preferential expression in cells of endothelial origin. We conclude that a nuclear factor with binding specificity for a GATA motif similar to that of the transcriptional activator GF-1 is necessary for the efficient and cell-specific expression of the human PPET-1 gene.

scholarly journals A nonerythroid GATA-binding protein is required for function of the human preproendothelin-1 promoter in endothelial cells.

1990 ◽  
Vol 10 (9) ◽  
pp. 4854-4862 ◽  
Author(s):  
D B Wilson ◽  
D M Dorfman ◽  
S H Orkin
Keyword(s):  
Growth Hormone ◽  
Endothelial Cells ◽  
Amino Acid ◽  
Dna Binding ◽  
Binding Protein ◽  
Cell Types ◽  
Dna Binding Protein ◽  
Core Motif ◽  
Transcription Activity ◽  
Shift Analysis

Endothelin-1 (ET-1) is a 21-amino-acid peptide synthesized by endothelial cells that has potent vasoconstrictor activity. Human ET-1 is derived from a 212-amino-acid prepropeptide, termed preproendothelin-1 (PPET-1). To identify cis-acting sequences essential for PPET-1 gene transcription, bovine aortic endothelial (BAE) cells were transfected with plasmids containing 5'-flanking sequences of the human PPET-1 gene fused to the human growth hormone gene as a reporter. Deletional analysis of these fusion plasmids showed that the sequence spanning positions -141 to -127 of the human PPET-1 promoter is required for full transcription activity. Introduction of clustered point mutations into this region of the promoter reduced transcription activity. Gel shift analysis, methylation interference, protein-DNA cross-linking, and oligonucleotide competition studies revealed that BAE cell nuclear extract contains a 47-kilodalton DNA-binding protein recognizing the core motif TATC (GATA) located at positions -135 to -132 of the PPET-1 promoter. The size and specificity of this DNA-binding protein resemble GF-1, a previously described transcription factor of erythroid cells that binds to the same core motif. Gel shift analysis indicated that GF-1 and the DNA-binding protein interacting with the PPET-1 promoter have different tissue distributions; the former is restricted to a subset of hematopoietic cells, and the latter is found in various cell types, including BAE, NIH 3T3, and HeLa cells. By using an antiserum to the C-terminal region of GF-1, the two proteins were also found to be antigenically distinct. When a growth hormone fusion plasmid containing the proximal 141 nucleotides of the PPET-1 promoter was transfected into a variety of cell types, these was preferential expression in cells of endothelial origin. We conclude that a nuclear factor with binding specificity for a GATA motif similar to that of the transcriptional activator GF-1 is necessary for the efficient and cell-specific expression of the human PPET-1 gene.

The DNA-binding protein II from Zymomonas mobilis. Complete amino acid sequence and interaction with DNA

Biochimie ◽  
1998 ◽  
Vol 80 (2) ◽  
pp. 109-116
Author(s):  
B. Laine ◽  
F. Chartier ◽  
F. Culard ◽  
D. Bélaïche ◽  
P. Sautière
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Amino Acid Sequence ◽  
Zymomonas Mobilis ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Complete Amino Acid Sequence

scholarly journals The gene G13 in the class III region of the human MHC encodes a potential DNA-binding protein

1996 ◽  
Vol 319 (1) ◽  
pp. 81-89 ◽  
Author(s):  
Anu KHANNA ◽  
R. Duncan CAMPBELL
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Leucine Zipper ◽  
Binding Protein ◽  
Coiled Coil ◽  
Single Copy ◽  
Dna Binding Protein ◽  
Lymphoid Tissues ◽  
Class Iii ◽  
Cell Extracts

G13 is a single-copy gene lying approx. 75 kb centromeric of the complement gene cluster in the class III region of the human MHC. The gene spans approx. 17 kb of DNA and has been shown to encode mRNA of approx. 2.7 kb that is present in cell lines representing lymphoid and non-lymphoid tissues, indicating that it is ubiquitously expressed. The complete nucleotide sequence of the 2.7 kb mRNA has been derived from cDNA and genomic clones. The longest open reading frame obtained for G13 codes for a 703 amino acid protein of approx. 77 kDa in molecular mass. Comparison of the putative G13 amino acid sequence with the protein databases revealed significant similarities with DNA-binding proteins of the leucine zipper class, including a human cAMP response element binding protein. G13 contains a bZIP motif, a region rich in basic amino acids adjacent to a coiled-coil leucine zipper domain, common to this class of proteins that is known to be involved in dimerization and DNA binding. Antibodies raised against a fragment encoding the C-terminal half of the putative G13 protein recognized a major polypeptide of approx. 86 kDa and a minor polypeptide of approx. 78 kDa on immunoblotting of U937 cell extracts; this has been confirmed by immunoprecipitation experiments. Even though it contained at least one potential bipartite nuclear localization signal, the G13 protein was present both in the cytoplasm and the nucleus of the fibroblast cells. Thus G13 might be a novel DNA-binding protein that is perhaps translocated to the nucleus in a regulated manner.

Mouse beta-globin DNA-binding protein B1 is identical to a proto-oncogene, the transcription factor Spi-1/PU.1, and is restricted in expression to hematopoietic cells and the testis

1993 ◽  
Vol 13 (5) ◽  
pp. 2929-2941
Author(s):  
D L Galson ◽  
J O Hensold ◽  
T R Bishop ◽  
M Schalling ◽  
A D D'Andrea ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Protein ◽  
Consensus Sequence ◽  
Cell Types ◽  
Dna Binding Protein ◽  
Globin Genes ◽  
Beta Globin ◽  
Chromosome 11 ◽  
Mobility Shift ◽  
Murine Erythroleukemia

The hematopoietic-specific DNA-binding protein B1 binds to the DNA consensus sequence AAAGRGGAARYG located twice in intervening sequence 2 of both of the mouse beta-globin genes (D. L. Galson and D.E. Housman, Mol. Cell. Biol. 8:381-392, 1988). B1 was cloned by expression of a murine erythroleukemia (MEL) cell cDNA library in transfected COS cells and screening by electrophoretic mobility shift analysis. B1 is identical to the proto-oncogene Spi-1/PU.1 (Spi-1), an ets family member. Protein-DNA contacts are shown to resemble those of the helix-turn-helix homeodomain proteins. By Northern (RNA) analysis, we found that Spi-1 mRNA is present at low levels during murine CFU-E maturation and is at least 20-fold higher in uninduced MEL, a transformed proerythroblast-like cell line which contains an activating/transforming insertion of spleen focus-forming virus at the Spi-1 locus. Dimethyl sulfoxide-induced MEL cell differentiation decreases Spi-1 mRNA to approximately 20% of the uninduced level before commitment occurs. In addition to erythroid cells, Spi-1 mRNA is present in B cells, myelomonocytes, and mast cells but not in T cells and nonhematopoietic cell types. In situ hybridization demonstrated Spi-1 mRNA expression in bone marrow, spleen, interstitial nonhepatocytes of the liver, and interstitial nontubular cells of the testis. The Spi-1 locus was mapped on human chromosome 11 to the same interval as ACP2 (lysosomal acid phosphatase), between the anonymous DNA markers D11S33 and D11S14. This region has not yet been found to be associated with a human malignancy.

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