Expression of a functional porcine growth hormone receptor cDNA in mouse L cells

Mol Cell Endocrinol. 1993 Jul;94(1):89-96. doi: 10.1016/0303-7207(93)90055-o.

Abstract

Porcine (p) growth hormone receptor (GHR) complementary DNA (cDNA) has been cloned and the primary amino acid structure was deduced from the nucleotide sequence. A comparison of pGHR to other GHRs revealed an approximately 70% similarity in amino acid sequence (Cioffi et al., 1990). Hybridization of this receptor cDNA to RNA samples isolated from various porcine tissues revealed a single RNA band of 4.2 kb. The full-length pGHR cDNA was subcloned into an eukaryotic expression vector, transcription of which was directed by the mouse metallothionein-I transcriptional regulatory sequence. Stable mouse L cell lines which express the pGHR cDNA were established. Approximately 80% of the cell lines were found to possess pGHR mRNA (approximately 2 kb) which corresponds to the length of the cloned pGHR cDNA. Binding studies showed that the stable cell lines were capable of specifically binding 125I-labeled pGH with a dissociation constant (Kd) of approximately 1.0 nM. The apparent molecular mass of the receptor, as determined by cross-linking studies, was found to be 118 kDa. Also, the receptor-ligand complex could be internalized. These results suggest that an active form of pGHR had been cloned and stably expressed in mouse L cells.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding, Competitive
  • Cloning, Molecular
  • DNA / genetics
  • Genetic Vectors
  • Growth Hormone / metabolism
  • L Cells / metabolism*
  • Mice
  • Molecular Sequence Data
  • Plasmids
  • RNA, Messenger / genetics
  • Receptors, Somatotropin / biosynthesis*
  • Receptors, Somatotropin / genetics
  • Recombinant Fusion Proteins / biosynthesis*
  • Recombinant Fusion Proteins / genetics
  • Swine / genetics

Substances

  • RNA, Messenger
  • Receptors, Somatotropin
  • Recombinant Fusion Proteins
  • Growth Hormone
  • DNA