*2.6. Subcellular Localization and Gene Ontology (GO) Analysis*

We predicted the subcellular localization of each protein and GO categories in each class for the evolutionary classification (Figure 4, Supplementary Table S7). Specific GO categories included epigenetic regulation of gene expression, transcriptional regulation, and organogenesis or organ morphogenesis (Figure 4). We confirmed the evolutionary trends of proteins with specific GO categories and their subcellular localization and found that 129 and 48 proteins in Classes 1–4 were expressed in the nucleus only or the nucleus and cytoplasm, respectively. Proteins in Classes 1–4 were represented in the epigenetic regulation of gene expression category, whereas transcriptional regulation was observed only in Classes 1 and 2, and organogenesis and organ morphogenesis were mainly observed in Class 2 (Figure 4).

**Figure 4.** Subcellular localization and specific GO categories of human RTT-related proteins: Phylogenetic trees show interactors, subcellular localization, and specific GO categories for each protein. The vertical axis shows 240 RTT-related proteins, and each bar shows MeCP2-interactor (red), CDKL5-interactor (green), and FOXG1-interactor (blue) (a1 and a2); cellular localization (b); epigenetic regulation of gene expression (c1); transcriptional regulation (c2); and organogenesis (c3).

### *2.7. Tissue and Organ Localization*

Tissue and organ expression data for 237 proteins were extracted from The Human Protein Atlas as transcripts per million (TPM) values [46]. In addition, four proteins were not expressed in the cerebral cortex. Tissues and organs with specific expression were identified using 195 RTT-related human proteins as queries (Figure S2, Supplementary Table S8). There were nine proteins that were specifically expressed in the cerebral cortex including apolipoprotein E, CDKL5, SATB2, spalt-like transcription factor (SALL)1, zinc finger protein (ZNF)483, FOXG1, (sex-determining region Y)-box (SOX)2, homeodomain-interacting protein kinase (HIPK)2, and histone cluster 2 H3 family member A.
