The role of RNA m6A methylation in insulin resistance in adipocytes

Abstract:

Objective To explore the role of RNA m⁶A methylation in insulin resistance in adipocytes.Methods We collected redundant subcutaneous adipose tissue samples from patients with type 2 diabetes and patients without type 2 diabetes to measure the RNA m⁶A modification level. A insulin resistance（IR） model was established by feeding C57BL/6J mice with a 60% high-fat diet for 16 weeks（n=5），while the control group was fed with a 10% low-fat diet for 16 weeks（n=5）. After successful modeling，the adipose tissue around the epididymis was taken to detect m⁶A methylation using epitranscriptomic microarrays. The changes in insulin signaling-related genes were determined by MeRIP-qPCR assay，RT-qPCR，and RIP assay. The effects of the small-molecule inhibitor STM2457 targeting methyltransferase like 3（METTL3） on insulin signaling-related genes in mice feeding a high-fat diet were investigated.Results The overall m⁶A modification levels were significantly increased in the adipose tissue of patients with type 2 diabetes and IR mice（patients 200 ng RNA，t=-8.375，P<0.001；patients 100 ng RNA，t=-3.722，P=0.006；patients 50 ng RNA，t=-4.937，P=0.001；mice 100 ng RNA，t=-3.590，P=0.023；mice 50 ng RNA，t=-2.760，P=0.025）. The epitranscriptomic assay detected high m⁶A methylation levels in 1 175 genes and low m⁶A methylation levels in 55 genes；182 genes showed significantly high m⁶A modification and low expression，including five key insulin signaling-related genes（AKT2，INSR，PIK3R1，ACACA，and SREBF1）. Direct binding between AKT2，INSR，ACACA，and SREBF1 and METTL3 was validated，and their m⁶A modification levels were positively regulated by METTL3. STM2457 significantly increased insulin sensitivity，and significantly upregulated the transcriptional levels of AKT2，INSR，ACACA，and SREBF1，suggesting a significant improvement in IR phenotype.Conclusion High-fat diet induces IR through METTL3，which mediates m⁶A hypermethylation of AKT2，INSR，ACACA，and SREBF1 to downregulate their expression and block insulin signaling in adipocytes.