FTO‐mediated m 6 A modification of SOCS1 mRNA promotes the progression of diabetic kidney disease

Abstract

Dear Editor, N 6-methyladenosine (m 6 A) is the most prominent and frequent internal messenger RNA (mRNA) modification and plays diverse roles in regulating functions of modified transcripts. 1 However, the role of m 6 A modification in kidney disease remains rarely understood, especially at the onset of diabetic kidney disease (DKD). 2 Here, we delineate the biological role of FTO-mediated m 6 A modification in DKD through imaging mass cytometry (IMC), LC/MS, and RNASeq methods. The results show that the loss of m 6 A levels by overexpressing FTO recapitulated human DKD by increasing the expression of suppressors of cytokine signalling 1 (SOCS1) protein level to alleviate inflammation response and kidney injury. Thus, FTO maybe a potential therapeutic target for DKD patients. To quantify the m 6 A level at cellular and spatial levels , we designed an IMC panel specific to kidney histology and used this to analyse kidney biopsies (Figure 1A and Table S1). IMC integrates IHC using metal isotope-tagged antibodies with laser ablation and mass-spectrometry-based detection to produce high-dimensional images, 3 which allow simultaneously quantified the m 6 A levels and its regulators. We identified 17 676 cells and quantified the levels of m 6 A, regulators, and spatial characteristics at single-cell level (Figures 1 and S1). We identified five dominant cell clusters of proximal tubules, distal convo-luted tubule, glomerulus (Glom) endothelial, macrophage, and stromal cell populations. 4 IMC and IHC data demonstrated that the m 6 A levels were significantly increased in several types of cells, and FTO expression was significantly reduced (Figure 2). The results suggest that FTO plays important roles in m 6 A dynamics in DKD. Further analysis revealed that serum m 6 A levels were significantly upregulated in T2D and DKD patients (Figures S2 and S3a). RNASeq data showed that FTO was significantly downregulated in T2D and DKD patients, whereas other regulators remained unchanged (Figures S3b and S4). Further data also showed that the m 6 A This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. levels remained significantly upregulated in T2D and DKD patients, and FTO level was negatively correlated with m 6 A levels (Figure S3c-e). Furthermore, re-analyses of public datasets revealed that FTO was significantly decreased in DKD or uremina patients (Figure S3f-h). To further investigate the role of FTO in DKD pathogenesis, high-concentration glucose (HG) treatment was used to simulate the phenotypes in DKD. 5 HG treatment significantly reduced FTO expression, whereas the m 6 A RNA level was significantly increased (Figure S5a-d). Moreover , other regulators remained unchanged after the HG treatment (Figure S6). FTO overexpression or knock-down significantly reduced or augmented the m 6 A levels, respectively (Figure S5e-h). HG often triggers glucose-response transcriptional factor ChREBP expression, 6 further analysis revealed that the FTO promoter had several ChREBP-binding sites, indicating HG may suppress FTO expression via activating ChREBP (Figure S5i). Together, these data reveal that the m 6 A modification levels are increased due to FTO downregulation in DKD. To investigate the molecular mechanism by how dysreg-ulated FTO is involved in DKD. We performed MeRIP-seq in HMC after FTO overexpression, and the results showed that m 6 A peaks were significantly enriched at the 3′-UTR region (Figure 3A,C) and were characterized by the RAACH motif (Figure 3B). Overall, 25 genes were affected at both RNA expression and m 6 A levels. Pathway enrichment analyses revealed that the genes affected are involved in inflammation (Figure 3D,E and Tables S6 and S7). More specifically, the m 6 A level of SOCS1, a key regulator of inflammation, was significantly reduced after FTO overex-pression (Figure 3F), which was confirmed by re-analyses of public datasets (Figure S7a). MeRIP-qPCR results showed that SOCS1 m 6 A level was significantly reduced on FTO overexpression (Figure 3G). Further data revealed that SOCS1 protein level was significantly increased when FTO was overexpressed (Figure 3H). SOCS1 mRNA expression was also positively associated with FTO expression Clin. Transl. Med. 2022;12:e942. wileyonlinelibrary.com/journal/ctm2 1 of 7 https://doi.