ESPE Abstracts

Background: GNAS (Guanine Nucleotide-Binding Protein, Alpha Stimulating) is an imprinted gene that encodes the alpha subunit of the stimulatory G protein (G_sa), which mediates the signaling of various G protein-coupled receptors. Inactivating genetic and epigenetic changes in GNAS, leading to G_sa deficiency, are associated with different subtypes of pseudohypoparathyroidism, which may include severe, early-onset obesity as part of their clinical spectra. More recently, G_sa deficiency has been linked to isolated, severe, early-onset obesity, suggesting that it may be an underappreciated cause of monogenic, non-syndromic obesity (Grüters-Kieslich et al., 2017; Mendes de Oliveira et al., 2021). The present study was conducted to investigate GNAS variations as potential causes of monogenic obesity in the population of Qatar.

Methods: Paediatric patients presenting to the endocrinology clinic in Sidra Medicine with unexplained, severe, early-onset obesity, their parents, and some of their siblings were tested for GNAS sequence and copy number variations using a gene panel and/or whole-genome sequencing. To functionally characterize the identified variants, we developed the first zebrafish model of G_sa deficiency-associated early-onset obesity by Morpholino-mediated knockdown of the ortholog, gnas. Moreover, we assessed the effects of two of the identified variants on the expression of GNAS and four lipid metabolism-associated genes in patient-derived T cells using quantitative reverse transcription PCR.

Results: We identified seven GNAS variants of interest among eight families, of which five were missense, one was a novel nonsense variant, and another was a small in-frame insertion. The nonsense variant and a missense one were classified as likely pathogenic, while the rest were variants of uncertain significance (VUSs), according to the ACMG/AMP guidelines. The morphant zebrafish larvae displayed an obese phenotype characterized by significantly enlarged yolk sacs, substantially increased neutral lipid accumulation, and considerably reduced metabolic rates, among other developmental abnormalities resembling those seen in humans with G_sa deficiency. In addition, the nonsense variant showed a strong correlation with early-onset obesity and was associated with reduced GNAS expression and increased PPARG, CPT1A, and FASN expression in the subjects carrying the variant.

Conclusion: Overall, these findings confirm the important role of loss of GNAS function in the pathogenesis of monogenic obesity. We successfully developed the first zebrafish model of G_sa deficiency-induced early-onset obesity by gnas knockdown using Morpholino technology. This work lays the foundation for the functional characterization of novel or previously reported GNAS VUSs using the zebrafish model.