molecular mechanism of abnormal growth hormone action

(1)在我们的患者中发现了GH-1基因的杂合错义突变R77 C。该突变体GH不仅不能单独刺激酪氨酸磷酸化，而且还抑制了野生型GH作为拮抗剂的活性。尽管患者的等位基因中存在基因组突变，但患者父亲的血清中为何不表达突变型GH，仍存在未解决的问题。为了排除基因组遗传的可能性，我们对其外祖母的GH-1基因进行了基因型调查。由于没有突变，体细胞嵌合体可能是更有可能的原因。(2)我们报道了一种新的杂合供体剪接位点突变的内含子9的GH受体（GHR）基因的同胞谁表现出部分GH不敏感和高血清GH结合蛋白（GHBP）水平。该突变导致剪接异常并产生由277个氨基酸组成的截短的GHR（GHR-277），其缺乏GHR的大部分胞内结构域，包括框1和框2。在本研究中，我们已经在体外表征了GHR-277在COS-7或CHO细胞中表达的功能。Scatchard分析显示，与野生型全长GHR（GHR-fl）相比，GHR-277具有两倍数量的结合位点。表达GHR-277的细胞的培养基中的GHBP水平比表达GHR-fl的细胞中的GHBP水平高约3倍。GHR-277和GHR-f1共转染时，GHR-277对GH诱导的STAT 5酪氨酸磷酸化起显性负效应。这些体外结果将很好地解释我们患者的临床特征。(3)在1例疑似生长激素不敏感综合征患者中，我们发现了一个杂合错义突变C422 F，但对该突变GHR的功能研究并不支持该突变是该患者生长障碍的原因。

(1) A heterozygous missense mutation R77C in the GH-1 gene was identified in our patient. This mutant GH not only failed to stimulate tyrosine phosphorylation by itself, but also inhibited the activity of the wild-type GH as an antagonist. There are still unresolved problems why the patient's father did not express the mutant GH in his serum despite the presence of a genomic mutation in his allele. To exclude the possibility of genomic implinting, we investigated the genotype of GH-1 gene in his grand mother. Since there was no mutation, somatic mosaicism may be more likely cause.(2) We have reported a novel heterozygous donor splice site mutation in intron 9 of the GH receptor (GHR) gene in siblings who showed partial GH insensitivity and high serum GH-binding protein (GHBP) levels. This mutation caused the splicing abnormality and produced the truncated GHR consisting of 277 amino acids (GHR-277), which lacked most of the intracellular domain of GHR, including both box 1 and 2. in this study we have characterized the function of GHR-277 expression in COS-7 or CHO cells in vitro. Scatchard analysis revealed that GHR-277 possessed twice the number of binding sites compared to wild-type full-length GHR (GHR-fl). The GHBP level in culture medium of GHR-277-expressing cells was approximately 3 times higher than that in GHR-fl expressing cells. The ligand induced internalization was significantly reduced comapred with that of GHR-fl. Moreover, in GH-induced tyrosine phosphorylation of STAT5, GHR-277 exerted a dominant negative effect when GHR-277 and GHR-fl were cotransfected. These in vitro results would well explain the clinical characteristics in our patients.(3) In a patient with a possible GH insensitivity syndrome, we found a heterozygous missense mutation C422F.However, functional study of this mutant GHR did not support this mutation as a cause of his growth failure.