Endocrine Abstracts

Self-limited delayed puberty is a condition that is frequently familial with strong genetic determinants. It has been linked to coding region sequence variation by next generation sequencing of affected individuals, identifying genetic regulation of gonadotropin-releasing hormone (GnRH) pathways underlying this condition. However, the role of epigenetic modifiers of human pubertal timing is underexplored. The hypothalamic-pituitary-gonadal axis is unique as it is active in three phases of life: foetal, infancy and then from puberty onwards. In-between these phases of life it is dormant. This is a process highly likely to be regulated by changes in DNA methylation. We analysed DNA methylation using the illumina EPIC array, for patients (n = 92) with delayed puberty who had no identified genetic cause for the condition, and related unaffected controls (n = 20). Quality control, annotation of CpG sites and differential methylation analysis was undertaken in R Studio. This revealed differentially methylated CpG sites between cases and controls linked to important genes known to play a role in the control of puberty and growth. Specifically, differentially methylated CpG sites were identified in both enhancer and promoter regions of these genes. Within the top 5 enhancer-associated CpG sites, two were linked to CADPS2 with q values 3.21x10^-19 and 7.31x10^-17 respectively. Key genes involved in upstream regulation of GnRH include TAC3, KMT2A and SIRT1 which showed increased methylation in CpG sites, within enhancer or promoter regions, in individuals with delayed puberty compared to controls. Genes associated with differentially methylated cytosines were then analysed to identify cellular pathways that were dysregulated. Over representation analysis identified multiple KEGG pathways previously associated with growth dysfunction as significantly altered. These included the cAMP pathway(p = 8.85x10^-08), the neuroactive ligand-receptor interaction pathway (p = 1.63x10^-05) and focal adhesion pathway (p = 1.63x10^-05). Our results suggest that changes in methylation of key regulatory genes contribute to the phenotype of self-limited delayed puberty.