Primary nucleotide structure of predominant and alternate splice forms of equine insulin-like growth factor I and their gene expression patterns in tissues.
OBJECTIVE
To isolate, clone, and determine primary nucleotide sequence of equine insulin-like growth factor I (IGF-I) and to examine IGF-I gene expression in tissues and cartilage from horses.


METHODS
Horses of various ages.


METHODS
Total RNA was isolated from tissues and purified. Complementary DNA (cDNA) was derived by reverse transcription and polymerase chain reaction (PCR) amplification and subcloned to plasmid vectors for sequencing and comparison with other species. Total RNA from various tissues was probed with radiolabeled cDNA or complimentary RNA constructs by use of northern blotting, tube hybridization, or PCR procedures to determine IGF-I expression patterns.


RESULTS
Nucleotide sequence of equine IGF-I was 90% homologous to that of cows, 88% homologous to that of humans and sheep, and 77% homologous to that of rats. Amino acid sequence was identical to that of humans, cows, dogs, and pigs. A larger PCR product (IGF-IB) was consistent with alternate splicing with retention of IGF-I exon 4 sequence, similar to rats and mice. Northern blot analysis revealed multiple IGF-I transcripts; predominant sizes were 1.6 and 4.5 kb. The IGF-I message was commonly detected in liver, kidney, and cartilage from young foals and was diminished in cartilage from a 12-month-old horse.


CONCLUSIONS
Nucleotide sequences of equine prepropeptides were different from those of other species, but the sequence coding the mature IGF-I peptide was more closely homologous. The larger IGF-IB form differed substantially in the carboxy-terminal. The biological action of the cleaved terminal was speculated to be autocrine feedback. Expression of IGF-I was apparent in many tissues, including cartilage, and was greater in immature horses.
Nixon
A J
AJ
Comparative Orthopaedics Laboratory, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853, USA.
Brower-Toland
B D
BD
Sandell
L J
LJ
D000375:Aging; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002356:Cartilage / Q000378:metabolism; D002417:Cattle; D017931:DNA Primers; D004285:Dogs; D015870:Gene Expression; D006736:Horses / Q000235:genetics*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009928:Organ Specificity; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013552:Swine
D017931:DNA Primers; D007334:Insulin-Like Growth Factor I