Cloning and characterization of an IGF-1 isoform expressed in skeletal muscle subjected to stretch

S Yang, M Alnaqeeb, H Simpson, G Goldspink

Research output: Contribution to journalArticlepeer-review

Abstract

To ascertain if IGF-1 is a regulator of local muscle growth, total RNA was extracted from rabbit muscle induced to undergo rapid hypertrophy using active stretch and from control muscles. This was analysed by Northern hybridization with a 280 base pair probe containing sequences derived from exons 3 and 4 of the insulin-like growth factor 1 gene. Two types of insulin-like growth factor 1 mRNA were shown to be strong expressed in the stretched muscles. In situ hybridization using the same probe (280 base pair) showed that IGF-1 is strongly expressed in muscle that is induced to grow rapidly and is expressed in the muscle fibres themselves. Using RT-PCR a single insulin-like growth factor 1 isoform cDNA (IGF-1Ea) could be cloned from the normal resting muscles. However, an additional isoform of insulin-like growth factor 1 (insulin-like growth factor 1Eb) was found to be expressed in stretched muscle undergoing hypertrophy. The E domain sequence of the additional isoform differs from the liver insulin-like growth factor 1Ea by the presence a 52 base pair insert. This changes the reading frame of the derived carboxyl-terminal resulting in a different precursor insulin-like growth factor 1 isoform. This insulin-like growth factor 1 mRNA probably encodes the precursor insulin-like growth factor 1 isoform that is responsible for local muscle growth regulation in response to mechanical stimulation. To confirm that alternative splicing of the insulin-like growth factor 1 gene occurs in muscle in response to physical activity, oligonucleotide primers were made which specifically amplify the cDNAs of two isoforms (insulin-like growth factors 1Ea and Eb) in the human as well as the rabbit. Following altered physical activity for 2 h to 6 days, appreciable levels of insulin-like growth factor 1Eb (in human the Ec) isoform were detected in skeletal muscle by using RT-PCR. In contrast very little if any of this splice variant could be detected in control muscle not subjected to stretch or extra physical activity.

Original languageEnglish
Pages (from-to)487-95
Number of pages9
JournalJournal of muscle research and cell motility
Volume17
Issue number4
Publication statusPublished - Aug 1996

Keywords

  • Animals
  • Blotting, Northern
  • Cloning, Molecular
  • Humans
  • Insulin-Like Growth Factor I
  • Muscle, Skeletal
  • Polymerase Chain Reaction
  • RNA Splicing
  • RNA, Messenger
  • Rabbits

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