The critical role of m6A methylation in the pathogenesis of Graves' ophthalmopathy

Abstract

Purpose To investigate the role of N6-methyladenosine (m6A) RNA modification in the pathogenesis of Graves' ophthalmopathy (GO). Methods Surgically excised extraocular muscles from 7 patients with GO and 5 subjects without GO were used. The global m6A levels in the specimens were determined using an m6A RNA methylation quantification kit. RNA sequencing (RNA-seq) was used to analyze the molecules involved in the regulation of m6A RNA methylation and the differential expression of mRNAs between the two groups (4 eyes, respectively). The expression of m6A RNA modification genes was evaluated by real-time PCR. The functional implications of the gene alterations between the GO and control specimens were determined by Gene Ontology analysis. Results The m6A level was significantly increased in the specimens of GO patients compared to the control specimens (P < 0.05). The expression of m6A methylation regulators, such as WT1 associated protein (WTAP), alkylation repair homolog protein 5 (ALKBH5), E74 like ETS transcription factor 3 (ELF3), YTH N6-methyladenosine RNA binding protein 2 (YTHDF2), YTHDF3 and YTH domain containing 2 (YTHDC2), was significantly upregulated (P < 0.05). Gene Ontology enrichment analysis showed that the most highly upregulated genes and biological pathways were related to the immune response and inflammatory processes such as lymphocyte activation, leukocyte differentiation, cytokine production and cytokine-mediated signaling pathways. Conclusions Our results suggest that m6A methylation may play a critical role in the pathogenesis of GO and that targeting genes that regulate m6A methylation may provide a new therapeutic approach for GO.

Potential Impact of N6-Methyladenosine RNA Methylation on Vision Function and the Pathological Processes of Ocular Diseases: New Discoveries and Future Perspectives

“…The expression of m6A writers (METTL1, METTL3, METTL14 and WTAP), erasers (FTO and ALKBH5) and readers (YTHDF1-3, YTHDC1 and 2, eIF3 and IGF2BP1-3) in ocular tissues has been reported in our previous study and other studies as well [11][12][13]15,39]. The expression of m6A modification factors (METTL3, METTL14, WTAP, FTO, ALKBH5, YTHDF1, YTHDF3 and YTHDC1) can be found in numerous ocular tissues (cornea, lens, retina, retinal pigmental epithelium [RPE], extraocular muscles and uveal melanoma cells) [41][42][43]45,51,[72][73][74][75][76]. m6A methylation usually varies among different tissues [77,78], indicating that m6A methylation is not only variable but also dynamic, and data obtained from one specific tissue or cell cannot represent the status of m6A methylation in other tissues or cells.…”

“…Of the ocular autoimmune diseases, Graves' ophthalmopathy (GO) is a typical autoimmune disorder that causes ophthalmopathy due to hyperthyroidism and excess autoantibodies production [255,258]. In a study of the role of m6A RNA modification in the pathogenesis of GO, we demonstrated that the expression of genes related to immune response and inflammatory processes such as lymphocyte activation and inflammatory pathways, which are associated with aberrant m6A modification factors, including m6A methylase WTAP, m6A demethylase ALKBH5, and m6A readers YTHDF2, YTHDF3, and YTHDC2, are significantly high in surgically excised extraocular muscles from patients with GO compared with that of normal subjects [45]. The results imply that m6A modification participates in the regulation of the pathogenesis of the autoimmune response in GO.…”

“…The aberrant expression of m6A modification factors has been shown to be related to many ocular diseases [42,45,51,[72][73][74][75]. The altered expression of m6A modification factors in ocular diseases has also been demonstrated.…”

“…Because RNA structural changes caused by the introduction of m6A may also alter the interaction between RNAs, proteins and chromatin, this new finding may extensively influence our comprehension of human diseases and their treatment [40], suggesting that m6A methylation regulates cellular functions and systemic diseases and is essential for vision function and eye diseases. Therefore, this review highlights the role of m6A Previous studies have demonstrated the expression of enzymes required for m6A modification in ocular tissues [41][42][43][44][45]. m6A modification and its biological functions require specific methyltransferases (writers), demethylases (erasers) and proteins that recognise methylated sites (readers) [46] (Fig.…”

“…A previous study demonstrated that NF-κB is listed among the top five of the 10 signalling pathways identified in the specimens of extraocular muscles of GO during the study of m6A methylation in the pathogenesis of GO. Notably, there is a close relationship between YTHDF expression and the NF-κB signalling pathway in controlling inflammation in the pathogenesis of GO [45]. The expression of NF-κB is regulated by the m6A methylation transferase METTL3 through its upstream gene MYD88, which controls cell differentiation [320], suggesting that RNA methylation, especially m6A modification plays an essential role in regulating signalling in many cells, including ocular cells.…”