Involvement of Sp1 in the transcriptional regulation of the rat insulin-like growth factor-1 gene.
Most insulin-like growth factor-I (IGF-I) transcripts are initiated in exon 1, but mechanisms of regulation are not well understood. Since potential Sp1 sites are found in footprinted regions within approximately 360 bp upstream and downstream from the major initiation sites in exon 1, we explored the involvement of Sp1 and Sp3 in regulation of IGF-1 expression. Gel shift assays showed strong Sp1 binding to the downstream site, but binding to the upstream site was weak; Sp1 bound to a CCTGCCCA sequence in downstream footprint region V, and Sp3 binding was centered on the same sequence. IGF-I basal promoter constructs containing a mutation in the downstream Sp1 site exhibited a 32% decrease in expression in CHO cells and a 75% decrease in HepG2 cells, indicating the importance of Sp1 for expression in vivo. Sp1 and Sp3 expression vectors provided three- to five-fold stimulation of wild-type IGF-I constructs, but had little effect on a construct containing a mutation in the downstream Sp1 site, and Sp1 had comparable effects in Drosophila SL2 cells. IGF-I heterologous promoter constructs exhibited similar responses: in both SL2 cells and CHO cells, stimulation by Sp1 was enhanced with constructs containing downstream region V. Since Sp1 also stimulated expression of concatamers of putative cis-acting sites fused to the SV40 promoter enhancer in pGL3, the results in combination indicate that the presence of IGF-I region V is sufficient to permit stimulation by Sp1.


CONCLUSIONS
Sp1 and related factors may play an important role in the regulation of IGF-I gene transcription, through interactions with region V downstream from the major initiation sites in exon 1.
Zhu
J L
JL
Department of Medicine, Emory University School of Medicine, Atlanta, GA 30322, USA.
Kaytor
E N
EN
Pao
C I
CI
Meng
X P
XP
Phillips
L S
LS
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005091:Exons; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D011401:Promoter Regions, Genetic; D051381:Rats; D016329:Sp1 Transcription Factor / Q000235:genetics*; D015533:Transcriptional Activation
D016329:Sp1 Transcription Factor; D007334:Insulin-Like Growth Factor I