Insulin-like growth factor-I regulates transcription of the elastin gene.
Neonatal rat aortic smooth muscle cell cultures were used to investigate the mechanisms by which insulin-like growth factor-I (IGF-I) up-regulates aortic elastogenesis. The addition of IGF-I (50 ng/ml) to quiescent smooth muscle cell cultures resulted in a 5-fold increase in the steady-state levels of tropoelastin mRNA beginning between 2 and 4 h and reaching maximal levels at 8 h. Addition of cycloheximide blocked the effect of IGF-I. Nuclear run-on transcription analyses of nuclei isolated from IGF-I-treated cells showed increased synthesis of new tropoelastin transcripts indicating that transcriptional activation is a major component of IGF-I up-regulation. Transient transfections with deletion constructs containing different portions of the elastin 5'-upstream region localized the IGF-I-responsive area to sequences between -195 and -136 base pairs and further showed that this region contains a negative element. Gel retardation assays using nuclear proteins extracted from control and IGF-I-treated cells demonstrated that IGF-I treatment results in the loss of binding complexes. Footprint analyses of specific binding complexes affected by IGF-I show the deprotection of two closely positioned sequences spanning positions -165 to -137 base pairs. These results suggest that IGF-I up-regulation of elastogenesis involves the abrogation of a negative element functionality.
Department of Biochemistry, Boston University School of Medicine, Massachusetts 02118.
B L
BL
Wolfe
C B
CB
Rich
H D
HD
Goud
A J
AJ
Terpstra
M
M
Bashir
J
J
Rosenbloom
G E
GE
Sonenshein
J A
JA
Foster
D000818:Animals; D000831:Animals, Newborn; D001011:Aorta / Q000378:metabolism*; D001483:Base Sequence; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D004247:DNA / Q000096:biosynthesis; D004549:Elastin / Q000096:biosynthesis* / Q000235:genetics*; D005786:Gene Expression Regulation; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013936:Thymidine / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014316:Tritium; D014333:Tropoelastin / Q000096:biosynthesis* / Q000235:genetics
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014333:Tropoelastin; D014316:Tritium; D007334:Insulin-Like Growth Factor I; D004247:DNA; D004549:Elastin; D015500:Chloramphenicol O-Acetyltransferase; D013936:Thymidine