Divergent RUNX2-Associated Regulation of FAM20A and FAM83H in the Enamel-Eruption Developmental Axis

Amelogenesis imperfecta (AI) associated with family with sequence similarity 20 member A (FAM20A) and family with sequence similarity 83 member H (FAM83H) may be accompanied by both enamel defects and tooth-eruption abnormalities; however, the tissue-level and molecular mechanisms underlying the relationship between these phenotypes remain poorly understood. To explore the mechanism, here we studied seven AI individuals from three families by oral and radiographic examination and whole-exome or whole-genome sequencing. Eruption abnormalities in FAM20A- and FAM83H-related AI were analyzed by integrating these seven cases with our previous retrospective datasets and newly reported cases from the literature. FAM20A and FAM83H expression was examined by RNA sequencing of human dental tissues and by immunohistochemistry in human dental follicle tissue and P0/P7 mouse tooth germs. RUNX2 regulation of FAM20A and FAM83H was assessed by CUT&Tag in dental follicle and RUNX2 siRNA knockdown. Genetic analysis identified two distinct heterozygous pathogenic truncating FAM83H variants in six patients from two families and compound heterozygous FAM20A variants in one patient from the third family. Eruption abnormalities were observed in the FAM20A patient and four of the six patients with FAM83H variants. In the combined dataset, eruption abnormalities were documented in 36 of 37 (97.3%) evaluable FAM20A cases and 18 of 58 (31.0%) evaluable FAM83H cases. In human dental follicle tissues surrounding impacted third molars, both proteins localized predominantly to epithelial lining components. In developing mouse tooth germs, both proteins were enriched in odontogenic epithelial/enamel-organ compartments with weaker staining in mesenchymal compartments. Dental-follicle RNA profiling further revealed an epithelial/keratin signature and detectable FAM20A/FAM83H expression. CUT&Tag identified RUNX2 binding at FAM20A but not FAM83H, and RUNX2 knockdown reduced FAM20A expression without a corresponding decrease in FAM83H. We conclude that FAM20A- and FAM83H-associated AI exhibit distinct patterns of eruption abnormalities in addition to enamel defects. Our findings support FAM20A as a candidate RUNX2-regulated target in dental follicle cells, whereas no comparable regulatory relationship was evident for FAM83H under the experimental conditions.