Title: Novel Variant Discovery in RORA in African Non-Syndromic Cleft Populations
Shaan Desai (Presenter)
University of Iowa College of Dentistry
TAMARA BUSCH, University of Iowa
James Park, University of Iowa College of Dentistry
Deepti Anand, University of Delaware
Carolina Bello, University of Puerto Rico School of Dental Medicine
Ada Rodriguez, University of Puerto Rico School of Dental Medicine
Cynthia Lo, University of Iowa
Muawia Mohamed, University of Iowa
Salil Lachke, University of Delaware
Azeez Butali, University of Iowa
Objectives: Orofacial clefts (OFCs) are the most common birth defects of the craniofacial region with a prevalence of approximately 1 in 700 live births. Multiple genes and environmental factors influence the risk of OFCs. Our objectives include identifying variants in the retinoid-related orphan receptor alpha gene (RORA) in non-syndromic clefts and testing variant function in silico. A recent genome-wide association meta-analysis reported a significant locus near chr15q22 for non-syndromic OFCs, and analysis using the SYSFACE bioinformatics tool identified RORA as the top candidate gene. RORA is an orphan nuclear receptor associated with many cellular activities, including cancer.
Methods: Two hundred eighty-eight saliva samples, obtained as part of a larger study of OFCs, from non-syndromic OFC populations from Ghana, Ethiopia, and Nigeria were used. We designed primers for all 12 exons of RORA and amplified the DNA samples via polymerase chain reaction. Samples were sent to Functional Biosciences for Sanger sequencing. We compared sequences to reference sequences to identify variants using UCSC BLAT (blast-like alignment tool) and Consed, a sequencing graphical editor. Any variants discovered were tested in silico with PolyPhen (Polymorphism Phenotyping) and SIFT (Sorting Intolerant From Tolerant) to determine their predicted degree of damage.
Results: A novel missense mutation was discovered in exon 1 of a Ghanaian. A guanine nucleotide was substituted for thymine. The affected individual is heterozygous at this locus: chr15:60,627,364. The affected codon changed from GGG to GTG. The resulting amino acid mutation was p.Gly4Val. The bioinformatics tool PolyPhen predicted the variant to be benign, whereas SIFT predicted it to be damaging.
Conclusions: We discovered a novel variant in one individual with non-syndromic cleft lip and palate. The variant is larger and more hydrophilic than the wild-type residue glycine, the most flexible amino acid. This flexibility may be necessary for the protein’s function; the mutation to valine may inhibit this function. This variant segregated in the family and was found in an unaffected sibling. Additional research to determine the functional role of this variant is required. Ultimately, we also hope to contribute to a more thorough understanding of the diverse etiology of OFCs.