Discovery of a novel deletion in PITX2 by dye-based quantitative PCR confirms that haploinsufficiency is a disease-causing mechanism for Axenfeld-Rieger Syndrome

Abstract

Purpose: FOXC1 and PITX2 mutations result in Axenfeld-Rieger syndrome (ARS), a genetically heterogeneous birth defect characterized by developmental defects of the anterior segment of the eye, an increased risk for glaucoma, and extraocular anomalies. We report the discovery and characterization of a novel PITX2 deletion in a small kindred with ARS.

Methods: Two familial patients (father and son) and one unrelated sporadic ARS patient were examined in the present study. Patient DNA samples were screened for FOXC1 and PITX2 mutations by sequencing, and for copy number variation by SYBR Green quantitative PCR analysis.

Results: DNA sequencing detected no sequence variations in the FOXC1 gene in any of the three ARS patients. However, a novel deletion involving the coding region of PITX2 was identified in the father-son pair. The novel deletion spans all known PITX2 exons as well as one upstream regulatory element. The son, of the father-son pair, additionally possesses a novel two-base-pair deletion in a non-coding exon of the remaining PITX2 allele. We also identified previously-recorded single nucleotide polymorphisms in non-coding regions of PITX2 in the cohort.

Conclusions: Our findings implicate a novel deletion of the PITX2 gene in the pathogenesis of ARS in an affected family. These results confirm that haploinsufficiency of PITX2 is a disease-causing mechanism for ARS. Furthermore, our results indicate that SYBR Green quantitative PCR is an efficient means of detecting causative PITX2 deletions in patients with ARS and may elevate the detection rate at this locus.

Keywords: 539 genetics • 537 gene screening • 421 anterior segment