DNA AND CELL BIOLOGY Volume 11, Number

1, 1992

Mary Ann Liebert, Inc., Publishers Pp. 43-50

Cloning and Characterization of the Human Insulin-Like Growth Factor-I Receptor Gene 5'-Flanking Region PAUL W. MAMULA* and IRA D.

GOLDFINE*>t

ABSTRACT The insulin-like growth factor-I receptor (IGFIR) is a membrane-bound glycoprotein that mediates the action of insulin-like growth factors. The cDNAs for the human IGFIR have been cloned and expressed, but the structures of the gene and its promoter have not been elucidated. In this study, we isolated an IGFIR promoter clone from a human chromosome 15 library. This clone contained the promoter, first exon, and a portion of the first intron. Sequence analysis of the 5' region that contained the promoter revealed that it lacked both TATA and CAAT boxes. The promoter contained binding sites for the transcription factors Spl, AP-2, and the epidermal growth factor receptor transcription factor (ETF). Primer extension analysis of IGFIR mRNA indicated the presence of a single transcription start site 1,012 bp upstream from the ATG. When the putative promoter was ligated into a promoterless CAT vector and transfected into HEPG2 cells, CAT activity was expressed, indicating that promoter activity was contained in this fragment. Other constructs containing the promoter and portions of the 5' untranslated region were used in transfection studies, and indicated that the 5' untranslated regions may play a role in promoter activity. Comparison of the human IGFIR promoter with that of the rat IGFIR promoter revealed significant sequence homology. Comparison of the IGFIR promoter with that of the human insulin receptor (IR) revealed structural similarities, although the arrangement of promoter elements differed.

INTRODUCTION factor-I receptor

Thehas been studied, its primary cDNA insulin-like growth

tains the intracellular

structure

tyrosine-specific kinase activity

(Froesch et ai, 1985). After IGF-I binds to its receptor, tyrosine kinase activity is activated, which leads to /3-subunit autophosphorylation and the initiation of an IGF-Ispecific signaling cascade (Jacobs et ai, 1983b; Ullrich et ai, 1986). While the IGFIR plays a major role in normal growth and differentiation and may play a role in human tumor biology (Rechler et ai, 1980; Roberts and LeRoith, 1988), little is

(IGFIR)

determined

expressed (Ullrich et (Ullrich et ai, 1986), and its ai, 1986; Steele-Perkins et ai, 1988; Kaleko et ai, 1990). This receptor is a membrane-bound glycoprotein consisting of two 135-kD a-subunits and two 95-kD subunits that are connected by disulfide bonds to form a functional »

33

GGGGCTCTTGTTTACCAGCATTAACTCCGCTGACCAAAAAAAAAGCGAAAAAACCCGAGCAGGAGCGAGCGCACCAGGCGAACTCGACACAGCGGGCAGAGCGAGACCGA

143

CGCCGCCAGCGAGCCTGCCCACCGCCCGCGGCTCCCCACCCCTCGGCCCGCTCCCCGCATCCCCCGCCCCCTCCACCCCCCTCCCGCGCGGGGGCAGCTCCACCCGCGCC

253

TCGCCTCGGCTGTGACCTTCAGCGACCCGGAGCCCCCGCGGCAGACCAGGCGGCGGCCCGCGGCGGCCGGGCGGGGGCCCGCGCGCGGCGGGCGCCGGCGCAGAGCCGGG

363

CGGCGCGGCGGGAGTGCTGAGCGCGGCGCGGCCGGCCCGCCGGCTTTGTGTGTGTCCTGGATTTGGGAAGGAGCTCGCCGCGGGGCCGCTGAGGGAGGAGGCGGCCGCGA

473

GGCGAGCCAGGAGGAGGAGGAGGAGGACCGGGAGCCGCTCATTCATTTTGACTCCGCGTTTCTGCCCCTCGCCGGCCTCGCCTGTCACCCGGACTTCGGGGCGATCTTGC

583

GAACTGCGTCGC&CCCTCCCGCGGCGGAAGCTCGGGCGTCCGGCCGCCTCCCGCGCGGCAGGGCCGGGCTTGTTTTTCCTCGCCTAGGCAGATTTCGGCTTTGCCCTTTC

693

TTTGCAGTTTTCCCCCCTTCCTGCCTCTCCGCGTTTGAAAATGGAGGCCGACGACGCCGACAGCCCGCCCGGCCGCCTCGGGTTCCCGACTCCGCCGAGCCCTGGGCCGC

803

Cloning and characterization of the human insulin-like growth factor-I receptor gene 5'-flanking region.

The insulin-like growth factor-I receptor (IGFIR) is a membrane-bound glycoprotein that mediates the action of insulin-like growth factors. The cDNAs ...
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