Rs1873147 Modulates Tpm1 To Increase Orofacial Cleft Risk Via Apoptosis

Rs1873147, a risk variant of non-syndromic cleft lip with or without cleft palate (NSCL/P) from a meta-analysis of genome-wide association studies. We aim to explore its regulatory mechanism in NSCL/P pathogenesis. The long-range regulatory role of rs1873147 was investigated through quantitative trait loci analysis (40 NSCL/P lip tissues), dual-luciferase reporter assay, zebrafish enhancer in vivo reporter assay, chromatin immunoprecipitation (ChIP), and chromosome conformation capture assay. We conducted dual-luciferase reporter assay, ChIP, electrophoretic mobility shift assay (EMSA), and supershift EMSA to identify the transcription factor binding to rs1873147 with allele-specificity. Functional impacts of rs1873147 mutation induced gene overexpression were tested in human embryonic cells and zebrafish models. Downstream molecules and pathways were identified through RNA sequencing, protein-protein interaction analysis, immunofluorescence co localization, and co-immunoprecipitation. The rescue effects of the key molecule were validated by cell phenotype experiments and TUNEL, acridine orange staining, and caspase-3 assays in zebrafish. Ethics approval was obtained from our institution. Rs1873147 is a well-documented susceptibility locus for NSCL/P, which promotes the affinity of transcription factor CEBEB through A>G mutation, remotely driving overexpression of TPM1. Zebrafish overexpression model showed malformations such as shortened body length, deformed ethmoid plate, and decreased iris pigment. Mechanistically, TPM1 interacts with ACTG1, activating cell apoptosis and increasing the risk of NSCL/P. This study updates the specific association mechanism between the 15q22 region and NSCL/P, providing guidance for the clinical prevention and treatment of NSCL/P.