Differential methylation of microRNA encoding genes may contribute to high myopia

Abstract

Introduction: High myopia (HM), an eye disorder with a refractive error ≤-6.0 diopters, has multifactorial etiology with environmental and genetic factors involved. Recent studies confirm the impact of alterations in DNA methylation and microRNAs (miRNAs) on myopia. Here, we studied the combined aspects evaluating to the role of methylation of miRNA encoding genes in HM. Materials and Methods: From the genome-wide DNA methylation data of 18 Polish children with HM and 18 matched controls, we retrieved differentially methylated CG dinucleotides localized in miRNA encoding genes. Putative target genes of the highest-ranked miRNAs were obtained from the miRDB and included in overrepresentation analyses in the ConsensusPathDB. Expression of target genes was assessed using the RNA sequencing data of retinal ARPE-19 cell line. Results: We identified differential methylation of CG dinucleotides in promoter regions of MIR3621, MIR34C, MIR423 (increased methylation level), and MIR1178, MIRLET7A2, MIR885, MIR548I3, MIR6854, MIR675, MIRLET7C, MIR99A (decreased methylation level) genes. Several targets of these miRNAs, e.g. GNAS, TRAM1, CTNNB1, EIF4B, TENM3 and RUNX were previously associated with myopia/HM/refractive error in Europeans in genome-wide association studies. Overrepresentation analyses of miRNAs' targets revealed enrichment in pathways/processes related to eye structure/function, such as axon guidance, transcription, focal adhesion, and signaling pathways of TGF-β, insulin, MAPK and EGF-EGFR. Conclusion: Differential methylation of indicated miRNA encoding genes might influence their expression and contribute to HM pathogenesis via disrupted regulation of transcription of miRNAs' target genes. Methylation of genes encoding miRNAs may be a new direction in research on both the mechanisms determining HM and non-invasive indicators in diagnostics.

Keywords: DNA methylation; childhood myopia; early-onset high myopia; epigenetic changes; miRNA encoding genes; microRNA target genes.

FIGURE 1

Detailed workflow of miRNA encoding genes methylation analyses in children with HM and matched controls. The research steps marked in white have been already published (Vishweswaraiah et al., 2019). Grey colour indicates the steps performed in the current study. TSS200 means 0–200 bases upstream of the transcriptional start site, TSS1500 means 200–1500 bases upstream of the transcriptional start site, UTR stands for untranslated region, FDR is false discovery rate, and SNP is single nucleotide polymorphism.

FIGURE 2

Comparisons of methylation levels of the highest-ranked CG dinucleotides between cases with HM and controls. Presented are CG dinucleotides with at least 10% methylation difference between HM cases and controls and location in miRNA encoding genes promoter regions. Standard deviation is included and asterisk (*) stands for the statistically significant difference in methylation level (FDR-corrected p-value < 0.05). (A) CG dinucleotides with increased methylation level in HM cases versus controls. (B) CG dinucleotides with decreased methylation level in HM cases versus controls.