In vivo footprinting and functional analysis of the human c-sis/PDGF B gene promoter provides evidence for two binding sites for transcriptional activators.
By in vivo DMS footprint and reporter gene analyses we identified two transcription factor binding sites in the human c-sis/PDGF B gene promoter. The low basal activity of the PDGF B promoter in HeLa and undifferentiated K562 cells, which express low PDGF B mRNA levels, and in PC3 cells, which express a high PDGF B mRNA level, results from binding of a weak transcriptional activator between positions -64 and -61 relative to the transcription start site. Cytotrophoblast-like JEG-3 cells, which do not express the 3.5 kb PDGF B mRNA, contain a transcriptional activator directed at the -64/-61 sequence, but DNA methylation may render the endogenous promoter inaccessible to this activator. A CCACCCAC element at position -61/-54 was identified as the in vivo binding site for a strong transcriptional activator in phorbol ester-treated megakaryocytic K562 cells, which express a high PDGF B mRNA level. Primary human fibroblasts, which do not transcribe the PDGF B gene, contain a transcriptional activator that recognizes an element between positions -60 and -45 but does not bind to the endogenous unmethylated promoter. Our results show that the complex expression pattern of the human PDGF B gene involves the cell type-specific expression of weak and strong transcriptional activators and regulation of promoter accessibility to these factors.
Dirks
R P
RP
Department of Biochemistry, University of Nijmegen, The Netherlands.
Jansen
H J
HJ
van Gerven
B
B
Onnekink
C
C
Bloemers
H P
HP
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D002874:Chromosome Mapping; D004247:DNA / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D008745:Methylation; D008969:Molecular Sequence Data; D010982:Platelet-Derived Growth Factor / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D015534:Trans-Activators / Q000378:metabolism*
D017931:DNA Primers; D010982:Platelet-Derived Growth Factor; D015534:Trans-Activators; D004247:DNA