Endocrine Abstracts

Background: Growth Hormone Insensitivity is usually caused by mutations in the Growth Hormone receptor (GHR). Patients present with short stature, high GH levels, low IGF-I levels and typical Laron syndrome facial features. Our centre previously described the first GHR pseudoexon mutation (42700896A>G, c.618+792A>G). The inclusion of this pseudoexon is predicted to cause in-frame insertion of 36 amino acid residues between exons 6 and 7. This insertion in the dimerization domain of the GHR results in defective trafficking rather than impaired signalling, causing a partial loss-of-function. As such, moderate postnatal growth failure is observed (Height SDS −3.3 to −6.0).

Hypothesis: Pseudoexons outnumber exons by 10-1 and variants in them may be a major contributor to disease burden.

Methods: We designed a custom short stature gene panel that interrogates both coding and non-coding regions to uncover such mutations. In-vitro splicing assays were performed using an exon trap vector (pET01, MoBiTec GmbH, Germany).

Results: We identified a homozygous GHR variant (42700940T>G, c.618+836T>G) in an Italian patient with classical Laron phenotype, severe postnatal growth failure and height SDS −7.5. Both unaffected, non-consanguineous parents were heterozygous for the mutation. This mutation is 44bp downstream of the original pseudoexon mutation and predicted in silico to create a donor splice site. Splicing analysis of this variant confirmed inclusion of a 152bp mutant pseudoexon in all transcripts with no evidence of normal splicing in contrast to the wild-type pseudoexon which showed no such inclusion. Inclusion of the pseudoexon will lead to a frameshift and premature truncation of the mRNA.

Discussion: This novel pseudoexon inclusion event will result in a truncated message which will either be destroyed by nonsense mediated mRNA decay or will lead to a truncated protein lacking the transmembrane and intracellular domains responsible for anchoring the protein in the membrane and signalling respectively. Given the undetectable GHBP levels in this patient, the former effect is suspected. This mutation will cause complete loss-of-function, consistent with the severe growth failure observed. This discovery highlights the potential for such splicing events to be more commonly causal for this and other rare diseases.