The effect of cellular density on insulin-like growth factor I (IGF-I) gene expression was characterized in several tumor-derived cell lines. IGF-I messenger RNA (mRNA) transcripts increased more than 200-fold when C6 glioma cells grew to postconfluence. IGF-I receptor and β-actin mRNAs were induced by 6- and 2-fold, respectively, as a function of confluence. IGF-I mRNA transcripts in GH3 and SK-N-MC cells increased about 4- to 5-fold in confluent cultures compared with sparse cultures. In OVCAR-3 cells, the IGF-I mRNA level remained constant as the cell density increased. Transient transfection experiments were performed with IGF-I exon 1 promoter/luciferase fusion constructs in C6 cells. The luciferase activity of a construct containing exon 1 sequence between +75 and +282 (the most 5′ transcription initiation site was designated +1) was stimulated by 2.5-fold in dense cultures compared with that in sparse cultures of C6 cells. Luciferase activities of other constructs containing at least 282 bp of exon 1 sequence were also stimulated about 2- to 4-fold by cell density. However, 3′ deletion to +192 led to loss of the cell density stimulatory effect. In contrast, luciferase activities of IGF-I promoter constructs were not altered by cell density in SK-N-MC cells. When the conditioned medium of low density C6 cultures was exchanged with that of high density cultures, the IGF-I mRNA level remained the same. In summary, cell density has a cell type- and gene type-specific effect on IGF-I gene expression. A cell density response element(s) may be located between +192 and +282 of the exon 1 promoter region in C6 cells.
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