DLX4 is associated with orofacial clefting and abnormal jaw development

Area of Science:

• Genetics
• Developmental Biology
• Craniofacial Abnormalities

Background:

• Cleft lip and/or palate (CL/P) are common congenital structural birth defects.
• The genetic underpinnings of many CL/P cases remain elusive.
• Homeobox genes, including DLX family members, are crucial in embryonic development.

Purpose of the Study:

• To investigate the genetic cause of CL/P in a patient with bilateral CL/P and her son.
• To determine the functional role of the identified DLX4 gene variant.
• To explore the potential involvement of DLX4 in human CL/P pathogenesis.

Main Methods:

• Exome sequencing to identify genetic variants in affected individuals.
• Sanger sequencing for variant verification.
• In situ hybridization in murine models to assess gene expression patterns.
• Functional assays using cell lines (human and HeLa) and zebrafish (Danio rerio) models to evaluate the impact of the DLX4 variant.
• Bioinformatic prediction of variant deleteriousness.

Main Results:

• A novel single nucleotide deletion (c.546_546delG) in the Distal-less 4 (DLX4) gene was identified in the patient and her son, predicting a loss-of-function variant (p.Gln183Argfs*57).
• The DLX4 variant was absent from population databases and predicted to be deleterious.
• Functional studies indicated the mutant DLX4 could not activate regulatory elements, suggesting haploinsufficiency.
• In vitro experiments showed reduced DLX4 expression altered the expression of other DLX genes and BMP4.
• Zebrafish models with targeted dlx4b knockdown exhibited craniofacial abnormalities.

Conclusions:

• This study identifies a disease-causing mutation in the DLX4 gene associated with human CL/P.
• The findings establish DLX4 as a potential candidate gene for nonsyndromic CL/P.
• DLX4 plays a critical role in craniofacial development, and its disruption can lead to CL/P.