In Vitro and In Vivo Expression of the Human Growth Hormone Analog hGH R77C
The naturally occurring human growth hormone analog, hGH R77C, has been described as a GH antagonist. The objective of this study was to confirm this observation and to further characterize the mechanism by which this molecule antagonizes the actions of GH and to compare the potency of this antagonist to the known GH antagonists (bGH G119K and hGH G120K) developed in our laboratory. The engineered mammalian expression plasmid pIG-MET-hGH R77C, which contains the hGH R77C gene, the mouse metallothionein I (mMET) transcriptional regulatory element to direct gene expression and the human growth hormone polyadenylation signal for proper mRNA processing, was engineered and used to generate mouse L cell lines and transgenic mouse lines that express hGH R77C. The mouse L cell lines were characterized and hGH R77C was found to act as a growth hormone receptor (GHR) agonist as opposed to an antagonist. Microinjection yielded 4 mice that possessed integrated hGH R77C DNA. All 4 of these mice possessed a large phenotype, suggesting that hGH R77C acted as a GHR agonist in vivo as well. In addition to their large size, hGH R77C transgenic animals also possessed a "short hair" phenotype and the females were infertile. All three of these phenotypes are observed in transgenic mouse lines that express wild type hGH. In conclusion, although previously reported to be a GH receptor antagonist, hGH R77C has an agonistic effect when expressed in either stable mouse cell lines or in transgenic mice. These data suggest that hGH R77C is acting differently when expressed in mice as opposed to man. This is possibly due to differences between the human and mouse GHR. Alternately, since the hGH R77C transgene is driven by the mMET promoter and not the wild type hGH promoter, the transgene is being produced by all cell types and is not pituitary specific. Also, protein processing and/or folding differences of hGH R77C in our mouse system could be different than those in man. Any or all of these possibilities could account for the differences in observed activity.
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:growth hormone hgh r77c transgenic mice
Date of Publication:01/01/2001