Sp1 binding is critical for promoter assembly and activation of the MCP-1 gene by tumor necrosis factor.
The monocyte chemoattractant protein-1 gene (MCP-1) is induced by the inflammatory cytokine tumor necrosis factor through the coordinate assembly of an NF-kappaB-dependent distal regulatory region and a proximal region that has been suggested to bind Sp1 as well as other factors. To provide a genetic correlation for Sp1 activity in this system, a cell line homozygous for a targeted truncation of the Sp1 gene was derived and examined. We found that the lack of Sp1 binding activity resulted in the inability of both the distal and proximal regions to assemble in vivo even though the binding of NF-kappaB to distal region DNA was unaffected in vitro. We also found that Sp1 and NF-kappaB were the minimal mammalian transcription factors required for efficient activity when transfected into Drosophila Schneider cells. Additionally, Sp3 was able to compensate for Sp1 in the Drosophila tissue cell system but not in the Sp1(-/-) cell line suggesting that Sp1 usage is site-specific and is likely to depend on the context of the binding site. Together, these data provide genetic and biochemical proof for Sp1 in regulating the MCP-1 gene.
Ping
D
D
Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Boekhoudt
G
G
Zhang
F
F
Morris
A
A
Philipsen
S
S
Warren
S T
ST
Boss
J M
JM
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D018932:Chemokine CCL2 / Q000235:genetics*; D054357:Chemokine CXCL10; D019743:Chemokines, CXC / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000502:physiology*; D004330:Drosophila / Q000235:genetics; D005786:Gene Expression Regulation; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016328:NF-kappa B / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011494:Protein Kinases / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
D018932:Chemokine CCL2; D054357:Chemokine CXCL10; D019743:Chemokines, CXC; D004268:DNA-Binding Proteins; D016328:NF-kappa B; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D011494:Protein Kinases; C065999:Sp1 kinase