Regulation of transcription of the TATA-less human complement component C4 gene.
The 5'-sequences flanking the human complement component C4 genes (C4A and C4B) have been analyzed for their ability to direct expression of a reporter gene in cell lines that constitutively express or do not express C4. No difference in the level of reporter gene expression was detected in cells transfected with C4A- or C4B-specific constructs. A series of reporter constructs containing progressively truncated C4 promoter fragments transfected into the hepatocyte Hep G2 cell line, identified the sequence contained within the region -178 to -39 as that associated with maximal reporter gene expression. This region contains consensus binding motifs for nuclear factor 1 (-110 to -97), Sp1 (-57 to -49), and three basic helix-loop-helix (-137 to -132, -98 to -93, and -78 to -73)-like transcription factors. Electromobility shift assays and DNase I footprinting analysis showed specific DNA-protein interactions of the C4 promoter at the nuclear factor 1, two E box (-98 to -93 and -78 to -73), and Sp1 binding domains. Site-directed mutagenesis of the Sp1 binding site resulted in total abrogation of reporter gene expression and mutation of the E box (-78 to -73) resulted in a 8-fold reduction in expression. We conclude that the Sp1 binding site at position -57 to -49 is critical for accurately initiated, basal transcription of C4.
Vaishnaw
A K
AK
Division of Medicine, Imperial College School of Medicine, London, UK.
Mitchell
T J
TJ
Rose
S J
SJ
Walport
M J
MJ
Morley
B J
BJ
D001483:Base Sequence; D003181:Complement C4 / Q000235:genetics*; D005786:Gene Expression Regulation; D015894:Genome, Human; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016385:TATA Box; D014158:Transcription, Genetic
D003181:Complement C4