Mutation Affecting the C-terminal of MSX1 Causes Non-syndromic Tooth Agenesis

Objectives: To identify the genetic defect that causing familial tooth agenesis in a Japanese family and corroborate the genotype/phenotype correlation in vivo.
Methods: Clinical examination and pedigree were first performed, and then genomic DNA was isolated from saliva samples. Mutation analysis was performed by amplifying MSX1, PAX9, AXIN2 and EDA exons and sequencing the products. After identifying the mutation, wild-type and mutated MSX1 vector were transfected separately to COS7 cells to observe the protein expression, localization and half-life of the protein. To corroborate the genotype/phenotype correlation, the region of Msx1 affected by the mutation was targeted using CRISPR/Cas system in mice.
Results: A novel frameshift mutation in MSX1 (p.A282RfrX301) was identified in the affected members of the family, located in the upstream region of the highly conserved C-terminal domain (MH6). In vitro functional analyses demonstrated that nuclear localization of the mutant protein was not altered, although the expression and the half-life of the mutant protein were reduced. MH6 was efficiently disrupted by CRISPR/Cas system. E16.5 mouse embryos showed cleft palate and delayed tooth development compared with the wild-type.
Conclusions: The novel mutation affecting the MH6 of MSX1 caused non-syndromic tooth agenesis in Japanese family. In vivo experiment demonstrated that MH6 plays an important role in palatogenesis and odontogenesis.