Short stature explained by dimerization of human growth hormone induced by a p.C53S point mutation

A homozygous mutation in growth hormone 1 (GH1) was recently identified in an individual with growth failure. This mutation, c.705G>C causes the replacement of cysteine at position 53 of the 191 amino acid sequence of 22 kDa human GH (hGH) with serine (p.Cys53Ser). This hGH molecule (further referred to as GH-C53S) lacks the disulfide bond between p.Cys53 and p.Cys165, which is highly conserved among species. It has been previously reported that monomeric GH-C53S has reduced bioactivity compared with wild-type GH (GH-wt) because of its decreased ability to bind and activate the GH receptor in vitro. In this study, we discovered that the substitution of p.Cys53 in hGH significantly increased formation of hGH-dimers in pituitary cells. We expressed his-tagged hGH variants in the cytoplasm of genetically modified Rosetta gami B DE3 Escherichia coli cells, facilitating high yield production. We observed that the bioactivity of monomeric GH-C53S is 25.2% of that of wild-type GH and that dimeric GH-C53S-his has no significant bioactivity in cell proliferation assays. We also found that the expression of GH-C53S in pituitary cells deviates from that of GH-wt. GH-C53S was exclusively stained in the Golgi apparatus, and no secretory granules formed for this variant, impairing its stimulated release. In summary, the unpaired cysteine C165 in GH-C53S forms a disulfide bond linking two hGH molecules in pituitary cells. We conclude that the GH-C53S dimer is inactive and responsible for the growth failure in the affected individual.