Fig. 3.

PAX7 target gene signature is a robust hallmark of FSHD skeletal muscle. a Gene set enrichment analysis was performed separately for activated and repressed targets of PAX7 against the Molecular Signatures database (MSigDB)^{37, 38}. Gene sets in MSigDB are frequently partitioned into ‘‘genes upregulated in phenotype’’ and ‘‘genes downregulated in phenotype’’. If such opposing gene sets were inversely significantly enriched across activated and repressed PAX7 targets (as assessed by Fisher’s exact test), they were considered robust. A bar plot displays the −log₁₀ (enrichment p-value) for PAX7 robust gene sets. A horizontal green line denotes p = 0.05. UP and DOWN on the x-axis refers to the partition of labeled gene sets into genes positively and negatively associated with the gene set respectively. PAX7 targets are enriched for repression of a HYPOXIA gene set, which specifically describes genes upregulated and downregulated following over-expression of a constitutively active form of HIF1α. Another gene set affected by PAX7 repression is EZH2 target genes. b A box plot demonstrates that the PAX7 target gene signature derived from 311 upregulated target genes and 290 downregulated target genes, validates as a biomarker on the RNA-seq FSHD muscle biopsy data set published by Yao et al.¹⁰. The box represents the interquartile range (IQR), with the median indicated by a line. Whiskers denote min (1.5*IQR, max (observed value)). ‘‘o’’ represents data points greater than 1.5 IQR from the median, n = 15 FSHD and n = 8 control muscle biopsies. The two-tailed Wilcoxon U-test p-value is given. c A forest plot displays the results of meta-analysis of the discriminatory power of the PAX7 target gene signature across five published microarray FSHD muscle biopsy data sets (in total n = 82 FSHD and n = 82 control muscle biopsies). The differential scores (FSHD score minus control score) alongside 95% confidence intervals are provided. Our PAX7 target gene signature is a significant biomarker on every individual FSHD muscle biopsy data set, and is strongly significant on meta-analysis. For single studies, a two-tailed Wilcoxon U-test was performed to assess significance, while a Fisher’s combined test was employed for overall assessment: *denotes p < 0.05, **denotes p < 0.01 and ***denotes p < 0.001