A Novel <b><i>GHR</i></b> Intronic Variant, <b><i>c.266+83G>T</i></b>, Activates a Cryptic 5′ Splice Site Causing Severe GHR Deficiency and Classical GH Insensitivity Syndrome

Abstract

Background/Aims: Mutations in the human growth hormone receptor gene (GHR) are the most common cause of growth hormone insensitivity (GHI) syndrome and insulin-like growth factor (IGF-1) deficiency. The extracellular domain of GHR (encoded by exons 2-7 of the GHR gene) can be proteolytically cleaved to circulate as GH-binding protein (GHBP). Methods: We evaluated the cause of classical GHI (Laron) phenotypes in 3 siblings. Results: Two brothers (aged 16.5 and 14.9 years) and their half-brother (aged 11.3 years) presented with extreme short stature (height standard deviation score, SDS, of -7.05, -6.34 and -8.02, respectively). The parents were consanguineous and of normal stature. Serum GHBP levels of probands were undetectable and circulating IGF-1 and IGF-binding protein-3 were abnormally low, but GH concentrations were elevated. Molecular analysis of the GHR gene revealed homozygous deletion of exon 3, a common polymorphism, and a novel c.266+83G>T variant within intron 4 which generated a 5′ donor splice site. Splicing events from this cryptic 5′ donor site resulted in retention of 81 intronic nucleotides in the GHR mRNA. Long-term rhIGF-1 therapy combined with leuprolide depot increased height by +2 to +3 SDS. Conclusion: The c.266+83G>T is the second intronic GHR mutation identified that activates a cryptic 5′ donor splice site. The abnormal splicing event led to early protein termination and undetectable serum GHBP concentrations.

Growth Hormone Receptor Mutations Related to Individual Dwarfism

“…Truncated GHR protein resulting from exon 8 skipping was directly secreted out of the cell [ 65 ], and truncated GHR missing 184 amino acids and another truncated GHR lacking all but five amino acids of the cytoplasmic domain could not mediate any effects of GH, nor was it internalized [ 64 ]. Truncated GHR 1–277 and 1–279 variants led to a translational frame shift that introduced a stop codon three to four amino acids after the GHR TMD, leading to truncation of the entire cytoplasmic domain ( Table 4 ) [ 6 , 66 ]. Using rhGH or rhIGF-1 to treat the patients with these mutations, although serum IGFBP-3 was normalized or below normal, IGF-1 serum levels were only modestly increased.…”

“…Patients with growth hormone insensitivity and without mutations in the GHR gene coding region should be screened for mutations in the noncoding regions, such as an intronic GHR mutation within intron 4 (266 + 83 G > T), which generates a 5’ donor splice site to retain 81 intronic nucleotides in the GHR mRNA. The abnormal splicing event caused early protein termination and undetectable GHBP in the serum [ 66 ]. Intron 6 (A (−1) > G (−1)) substitution lead to the skipping of exon 6 and premature termination of the mRNA message ( Table 5 ) [ 12 , 44 ].…”