Correlations between phenotype and gene region-specific episignatures in Rubinstein-Taybi syndrome and Menke-Hennekam syndrome

Abstract Background: Rubinstein-Taybi syndrome (RSTS) and Menke-Hennekam syndrome (MKHK) are two rare Mendelian disorders presented with variable degrees of intellectual disability and different facial dysmorphism. They are caused by loss-of-function (LOF) variants or missense/inframe deletion variants in the exon 30 and 31 of the CREBBP gene respectively, which is involved in histone modification and chromatin remodeling. Genetic defects in numerous genes have been found to disrupt epigenomic profiles including DNA methylation (DNAm) patterns (referred as episignature) in affected individuals. To further investigate the mechanism of CREBBPrelated disorders, human induced pluripotent stem cells (hiPSCs) are applied to study the DNAm alteration. Results: We presented RSTS and MKHK individuals with distinct clinical features. Detailed phenotype analysis showed that RSTS patients with nonsense-mediated mRNA decay evasion (NMD-evasion) variants had atypical facial dysmorphism and severer medical problems compared to the classical RSTS caused by LOF CREBBP variants. MKHK patients with variants in intrinsically disordered region (IDR) showed resemblant features. Further investigations elucidated these clinical conditions in methylation change. Genome-wide DNAm analysis of 9 RSTS and 8 MKHK patients and 33 controls identified two specific peripheral blood episignatures: RSTS and MKHK_IDR compared to matched normal controls. Methylation alterations in RSTS cases with NMD-evasion variants were mildly different from that of classical RSTS. MKHK subjects with variants outside the IDR did not obey the MKHK_IDR episignature. By interrogating DNAm in hiPSCs of 5 RSTS, 4 MKHK compared with 12 controls, we observed hypermethylated DNAm profiles of RSTS and MKHK in embryonic stage. Different methylation regions (DMRs) overlapping genes in hiPSCs of RSTS and MKHK play a role in embryonic development and organogenesis. Furthermore, DNAm patterns for hiPSCs of RSTS and MKHK were enriched for genes relevant for multicellular organismal homeostasis or transcriptional binding. Conclusions: We identified the type and locus of variants in the CREBBP gene as responsible for the RSTS and MKHK episignatures, consistent with phenotype analysis. DNAm profile analysis of hiPSCs revealed meaningful biological processes associated with embryonic development.