Functional SNPs within the intron 1 of the PROP1 gene contribute to combined growth hormone deficiency (CPHD)

Context: Mutations within the PROP1 gene represent one of the main causes of familial combined pituitary hormone deficiency (CPHD). However, most of the cases are sporadic with an unknown genetic cause. Objective: The aim of this study was the search for low penetrance variations within and around a conserved regulatory element in the intron 1 of PROP1, contributing to a multifactorial form of the disease in sporadic patients. Methods and Patients: A fragment of 570 bp encompassing the conserved region was sequenced in 107 CPHD patients and 294 controls, and an association study was performed with the four identified variants, namely c.109+435G>A (rs73346254), c.109+463C>T (rs4498267), c.109+768C>G (rs4431364), and c.109+915-917ins/delTAG (rs148607624). The functional role of the associated polymorphisms was evaluated by luciferase reporter gene expression analyses and EMSA. Results: A statistically significant increased frequency was observed in the patients for rs73346254A (P = 5 × 10^-4) and rs148607624delTAG (P = 0.01) alleles. Among all the possible allele combinations, only the haplotype bearing both risk alleles showed a significantly higher frequency in the patients vs. controls (P = 4.7 × 10^-4) and conferred a carrier risk of 4.19 (P = 1.2 × 10^-4). This haplotype determined a significant decrease of the luciferase activity in comparison with a basal promoter and the other allelic combinations in GH4C and MCF7 cells (P = 4.6 × 10^-6; P = 5.5 × 10^-4, respectively). The EMSA showed a differential affinity for nuclear proteins for the alternative alleles of the two associated variations. Conclusions: Variations with a functional significance conferring susceptibility to CPHD have been identified in the PROP1 gene, indicating a multifactorial origin of this disorder in sporadic cases.