Prolonged retention after aggregation into secretory granules of human R183H-growth hormone (GH), a mutant that causes autosomal dominant GH deficiency type II.
Human R183H-GH causes autosomal dominant GH deficiency type II. Because we show here that the mutant hormone is fully bioactive, we have sought to locate an impairment in its progress through the secretory pathway as assessed by pulse chase experiments. Newly synthesized wild-type and R183H-GH were stable when expressed transiently in AtT20 cells, and both formed equivalent amounts of Lubrol-insoluble aggregates within 40 min after synthesis. There was no evidence for intermolecular disulfide bond formation in aggregates of wild-type hormone or the R183H mutant. Both wild-type and R183H-GH were packaged into secretory granules, assessed by the ability of 1 mM BaCl2 to stimulate release and by immunocytochemistry. The mutant differed from wild-type hormone in its retention in the cells after packaging into secretory granules; 50% more R183H-GH than wild-type aggregates were retained in AtT20 cells 120 min after synthesis, and stimulated release of R183H-GH or a mixture of R183H-GH and wild-type that had been retained in the cell was reduced. The longer retention of R183H-GH aggregates indicates that a single point mutation in a protein contained in secretory granules affects the rate of secretory granule release.
Zhu
Yong Lian
YL
Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Conway-Campbell
Becky
B
Waters
Michael J
MJ
Dannies
Priscilla S
PS
D017609:Barium Compounds / Q000494:pharmacology; D002460:Cell Line; D002712:Chlorides / Q000494:pharmacology; D004220:Disulfides / Q000378:metabolism; D004355:Drug Stability; D015870:Gene Expression; D019382:Human Growth Hormone / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D007553:Isotope Labeling; D009154:Mutation; D022142:Secretory Vesicles / Q000378:metabolism*; D012995:Solubility; D013462:Sulfur Radioisotopes; D014162:Transfection
D017609:Barium Compounds; D002712:Chlorides; D004220:Disulfides; D013462:Sulfur Radioisotopes; C024986:barium chloride; D019382:Human Growth Hormone