Additional file 1 of Multiomics profiling of DNA methylation, microRNA, and mRNA in skeletal muscle from monozygotic twin pairs discordant for type 2 diabetes identifies dysregulated genes controlling metabolism

Additional file 1: Supplemental Table 1 Clinical characteristics of donors of muscle cells. Supplemental Table 2 Genes contributing to the enrichment scores of significantly regulated pathways obtained from the gene set enrichment analysis in muscle tissue from twins with type 2 diabetes compared with co-twins without type 2 diabetes. Supplemental Table 3 Gene sets with differential expression in myoblasts from subjects with type 2 diabetes versus controls (gene set enrichment analysis with q < 0.05). Supplemental Table 4 Gene sets with differential expression in myotubes from subjects with type 2 diabetes versus controls (gene set enrichment analysis with q < 0.05). Supplemental Table 5 Differentially expressed genes between twins with type 2 diabetes and co-twins without diabetes (P < 0.05). Supplemental Fig. 1 (A) Differentially expressed genes between monozygotic (MZ) twins with type 2 diabetes (T2D) and co-twins without T2D based on nominal P values (P < 0.05). Log2 fold changes in the volcano plot are shown when comparing expression in muscle from MZ twins with versus co-twins without T2D. The dashed line indicates P < 0.05. Gene labels are based on the top 20 most significant transcripts, however of those, only 10 labels are displayed as there were no annotations for the other 10. (B) Overlap of genes with differential expression in the skeletal muscle from MZ twins discordant for T2D (P < 0.05), in skeletal muscle from 15 subjects with T2D or impaired glucose tolerance (IGT) versus 362 normal glucose tolerance (NGT) controls (q < 0.05), as well as in cultured myoblasts and myotubes from 13 T2D versus 13 NGT unrelated subjects (P < 0.05). These results are also presented in Supplemental Table 5. (C) Schematic summary, partly based on Fig. 1A, highlighting some key results from this study. Bold text marks findings, we could validate in muscle biopsies or cells from unrelated subjects and/or based on functional experiments overexpressing two miRNAs in cultured human myotubes. Supplemental Table 6 Differentially expressed genes in myoblasts from subjects with type 2 diabetes compared with controls among the 863 genes differentially expressed in muscle from the type 2 diabetes discordant twin pairs (P < 0.05). Supplemental Table 7 Differentially expressed genes in myotubes from subjects with type 2 diabetes compared with controls among the 863 genes differentially expressed in muscle from the type 2 diabetes discordant twin pairs (P < 0.05). Supplemental Table 8 JASPAR motif search results in the set of differentially expressed genes between twins with type 2 diabetes and co-twins without type 2 diabetes. Supplemental Table 9 (A) Differentially methylated CpG sites in muscle from twins with type 2 diabetes compared with co-twins without type 2 diabetes (P < 0.05). (B) Average DNA methylation levels in different gene and CpG island regions using Illumina’s annotations [18] in muscle from twins with T2D compared with non-diabetic co-twins. Supplemental Table 10 Differentially methylated CpG sites in myoblasts obtained from 14 subjects with type 2 diabetes compared with 14 unrelated controls among the 15,647 sites differentially methylated in muscle from the type 2 diabetes discordant twin pairs (P < 0.05). Supplemental Table 11 Differentially methylated CpG sites in myotubes obtained from 14 subjects with type 2 diabetes compared with 14 unrelated controls among the 15,647 sites differentially methylated in muscle from the type 2 diabetes discordant twin pairs (P < 0.05). Supplemental Table 12 Differentially expressed miRNAs in skeletal muscle from twins with type 2 diabetes compared with co-twins without type 2 diabetes (P < 0.05). miR family names from miRBase indicate the relatedness between the miRNAs. Families with more than one member are shown. Supplemental Table 13 Intra–twin-pair correlations between miRNA expression and 2-h glucose, fasting glucose levels and HbA1c (P < 0.05). Within–twin-pair differences of the measures were calculated by subtracting the value of the co-twin without type 2 diabetes from the value of the twin with type 2 diabetes and correlations were analyzed using Spearman statistics. Supplemental Table 14 This table presents 569 predicted targets of the 69 differentially expressed miRNAs presented in Supplementary Table 12 and which overlap with the 823 differentially expressed genes presented in Supplementary Table 5 using the multiMiR tool ( http://multimir.ucdenver.edu/ ) which integrates the following tools: miRTarBase ( https://mirtarbase.cuhk.edu.cn/~miRTarBase/miRTarBase_2022/php/index.php ), TarBase ( https://dianalab.e-ce.uth.gr/tarbasev9 ), DIANA-microT ( https://dianalab.e-ce.uth.gr/microt_webserver/#/ ), MicroCosm ( http://www.ebi.ac.uk/enright-srv/microcosm/ ), miRanda ( http://www.microrna.org/ ), miRDB ( https://mirdb.org/ ), PITA ( http://mirtoolsgallery.tech/mirtoolsgallery/node/1066 ), ELMO ( http://www.mirz.unibas.ch/ ), TargetScan ( https://www.targetscan.org/vert_80/ ), PicTar ( https://pictar.mdc-berlin.de/ ), and miRecords ( https://www.hsls.pitt.edu/obrc/index.php?page=URL1237998207 ). Supplemental Table 15 Intra–twin-pair correlations between differences in miRNA expression and methylation in/near the gene (P < 0.05). Within–twin-pair differences of the measures were calculated by subtracting the value of the co-twin without type 2 diabetes from the value of the twin with type 2 diabetes and correlations were analyzed using Spearman statistics.