GHR gene transcript heterogeneity may explain phenotypic variability in GHR pseudoexon (6Ψ) patients

Abstract

Objectives The homozygous GH receptor (GHR) pseudoexon (6Ψ) mutation leads to growth hormone insensitivity (GHI) with clinical and biochemical heterogeneity. We investigated whether transcript heterogeneity (6Ψ-GHR to WT-GHR transcript ratio) and/or concurrent defects in other short stature (SS) genes contribute to this. Methods 6Ψ-GHR and WT-GHR mRNA transcripts of four 6Ψ patients (height SDS −4.2 to −3.1) and one control fibroblast were investigated by RT-PCR. Transcripts were quantified by qRT-PCR and delta delta CT analysis and compared using ANOVA with Bonferroni correction. In eleven 6Ψ patients, 40 genes known to cause GHI/SS were analysed by targeted next generation sequencing. Results RT-PCR confirmed 6Ψ-GHR transcript in the 6Ψ patients but not in the control. 6Ψ-GHR transcript levels were comparable in patients 1 and 3 but significantly different among all other patients. The mean 6Ψ:WT transcript ratios ranged from 29–71:1 for patients 1–4 and correlated negatively with height SDS (R = −0.85; P < 0.001). Eight deleterious variants in six genes were detected, but the number of gene hits did not correlate with the degree of SS in individual 6Ψ patients. Conclusion Variable amounts of 6Ψ- and WT-GHR transcripts were identified in 6Ψ patients but no 6Ψ transcript was present in the control. Higher 6Ψ:WT-GHR transcript ratio correlated with SS severity and may explain the phenotypic variability. Analysis of known SS genes suggested that phenotypic variation is independent of the genetic background. This is the first report of transcript heterogeneity producing a spectrum of clinical phenotypes in different individuals harbouring an identical homozygous genetic mutation.

Growth Hormone Receptor (GHR) 6Ω Pseudoexon Activation: A Novel Cause of Severe Growth Hormone Insensitivity

“…This resulted in defective trafficking (and concomitant reduced cell surface expression) rather than impaired signaling, causing a partial loss of function ( 11 ). As such, moderate postnatal growth failure was observed (Height SDS -3.3 to -6.0) ( 14 ). The intronic 6Ψ GHR mutation, 792 bases into the intron, was identified using homozygosity mapping of several polymorphic markers surrounding the GHR ( 10 ).…”

“…Interestingly, only 50% of the 20 6Ψ patients had “classical” GHI facial features ( 13 ). This milder, very variable phenotype (even among affected members of the same family) may be explained by the efficiency of splicing events that result in GHR transcript variability, that is, the relative abundance of different GHR transcripts ( 10 , 14 ). We report a novel GHR 6Ω pseudoexon resulting in severe postnatal growth failure/classical Laron syndrome.…”

GH Resistance Is a Component of Idiopathic Short Stature: Implications for rhGH Therapy

“…The phenotype of 6y subjects is therefore variable, more so than in other extracellular GHR mutations and it has recently been reported that variable amounts of 6y-and wild type-GHR transcripts were identified in 6y patients. Higher 6y:wild-type GHR transcript ratio correlated with the severity of the short stature phenotype (35).…”