Patterns and rates of exonic de novo mutations in sporadic Hirschsprung disease patients

Abstract

Hirschsprung disease (HSCR, aganglionic megacolon; 1 in 5000 live births) is a disorder of the enteric nervous system (ENS) characterized by the absence of enteric neurons along a variable length of the intestine. HSCR most commonly presents sporadically, although it can be familial (5-20% of the patients). The sporadic form of the disorder is believed to be a genetically complex disease with both de novo and/or inherited genetic lesions. To assess the role of de novo mutations (DNM) in sporadic HSCR, we performed exome sequencing on 16 HSCR patients and their unaffected parents. Standard BWA/GATK pipeline was used to map the sequence reads to human reference genome 19 and call genomic variants for all 48 samples simultaneously. Exonic DNM mutations were identified using KGGSeq. DNM candidates were scrutinized using Integrative Genomics Viewer (IGV) and those variants deemed plausible were validated by Sanger sequencing. In total we confirmed 20 DNM mutations (17 SNVs, 3 Indels) in 16 genes. Five DNM were identified in RET (major HSCR gene). CCR2, COL6A3, MED26, NUP98, HMCN1 and DENND3 had DNM mutations and were found mutated in independent HSCR patients. Importantly, some of these genes are members of pathways involved in the development of the ENS and the encoded proteins interact with known key signaling molecules. The overall exonic DNM mutation rate is 1.25 per HSCR trio, with 10 out of 16 (62.5%) patients harboring ≥ 1 DNM mutations. Therefore, DNM mutations, as inherited mutations, contribute to the development of sporadic HSCR.