An alternatively spliced human insulin-like growth factor-I transcript with hepatic tissue expression that diverts away from the mitogenic IBE1 peptide.
An alternatively spliced transcript of the human insulin-like growth factor-I (IGF-I) gene is described. The transcript was identified in human liver RNA by reverse transcriptase-polymerase chain reaction, cloning, and sequencing. It contained IGF-I exons 3 and 4, 49 basepairs of exon 5, then exon 6 (exon 4-5-6). The 5'-donor site at the exon 5-6 junction was a cryptic 5'-donor splice site (IGF633). The 3'-acceptor site of the splice was the usual intron-exon 6 junction. A second pair of primers across the exon 5-exon 6 junction was used to confirm the presence of the transcript by reverse transcriptase-polymerase chain reaction. Cloning and sequencing this second fragment confirmed the presence of this splice in human liver. The exon 4-5-6 transcript was quantified at about 10% relative to the exon 4-6 transcript in human livers (n = 7 subjects), but was not detected in other tissues. The exon 4-5-6 transcript was found in cultured human hepatoma HepG2 cells and increased, relative to exon 4-6 transcripts, in response to GH, but not in cultured human lymphoblast IM-9 cells. The exon 4-5-6 splice predicts a prepro-IGF-I of 158 amino acid residues, with an E-peptide sequence of 24 residues (Ec). The deduced Ec peptide sequence is 73% homologous to the rat Eb-peptide sequence. The predicted final residues of the Ec peptide are frameshifted exon 6 codons ending in an in-frame stop codon. The predicted peptide sequences of Ec and Eb differ at the cleavage site of the Eb-peptide fragment (IBE1), which has been shown to have mitogenic activity. These data suggest that 1) the exon 4-5-6 splice has hepatic tissue expression and occurs by the use of a cryptic 5'-donor consensus splice site (IGF633) in exon 5; 2) exon 4-5-6 can be hormonally regulated in cultured human HepG2 cells; 3) exon 4-5-6 is the human counterpart of the rat IGF-IEb, because the complementary DNA and predicted sequences are homologous; and 4) the production of IBE1 is potentially regulated by alternative splicing.
Chew
S L
SL
Department of Endocrinology, St. Bartholomew's Hospital, London, United Kingdom.
Lavender
P
P
Clark
A J
AJ
Ross
R J
RJ
D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular; D002455:Cell Division; D002460:Cell Line; D017931:DNA Primers; D005091:Exons; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis* / Q000235:genetics*; D008099:Liver / Q000378:metabolism*; D008113:Liver Neoplasms; D008214:Lymphocytes; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011498:Protein Precursors / Q000096:biosynthesis; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D010446:Peptide Fragments; D011498:Protein Precursors; C093199:preproinsulin-like growth factor I; D007334:Insulin-Like Growth Factor I