Mitochondria-related gene and protein changes in craniosynostosis: integrated transcriptomics and Fgfr2C361Y/+ cranial suture proteomics

Craniosynostosis is defined by premature cranial suture fusion and is biologically heterogeneous. To map mitochondrial-associated signals in craniosynostosis and rank follow-up candidates, we integrated two public microarray datasets (GSE27976, GSE50796), corrected batch effects, and analyzed 14,186 shared genes using limma. This identified 798 nominal DEGs (388 upregulated and 410 downregulated), of which 19 remained significant after Benjamini–Hochberg correction. Intersecting the nominal DEG list with the MitoCarta 3.0 inventory yielded 24 mitochondrial DEGs (MitoDEGs). Complementary feature selection reduced these 24 MitoDEGs to an eight-gene panel (TMEM11, SLC25A21, GPT2, CYP27A1, MRPS30, ACAA2, GSR, and LIG3); a multigene score reached an apparent AUC of 0.806 in the integrated dataset. Correlation-based co-expression analyses linked the panel to mitochondrial translation, electron transport, amino-acid metabolism, redox control, and cell–matrix signaling. Among craniosynostosis cases, consensus clustering on the eight genes separated two molecular subtypes with distinct GSVA pathway profiles. For experimental support in a genetically defined mouse model, we profiled bilateral coronal suture complexes from Fgfr2C361Y/+ knock-in (KI) pups and WT littermates. Jess capillary immunoassay showed higher CYP27A1 abundance in KI sutures (P = 0.0276), whereas ACAA2, LIG3, MRPS30, and TMEM11 were not significant. Data-independent acquisition (DIA) proteomics identified 523 differentially abundant proteins (516 increased, 7 decreased in KI), followed by stricter-threshold reporting, sensitivity analysis, and threshold-free rank-based enrichment. MitoCarta proteins and mitochondrial pathways remained supported under these more conservative analyses. These results support mitochondria-associated transcriptomic and proteomic changes in craniosynostosis and prioritize a limited set of mitochondrial candidates for future mechanistic work.