Mowat Wilson syndrome (MWS) is a rare neurodevelopmental disorder caused by mostly heterozygous loss-of-function variants in ZEB2. Affected individuals show considerable wide variability in clinical presentation. In particular, Hirschsprung disease (HSCR) occurs in only a subset of patients, suggesting that additional genetic factors may modify disease penetrance. To investigate this possibility, we performed whole-genome sequencing of two parent-child trios in which the probands carried pathogenic de novo ZEB2 variants but differed in enteric phenotype: one individual with MWS and long-segment HSCR and another with MWS without HSCR. In both probands, the ZEB2 variants represent the primary causative genomic diagnosis, and no additional rare coding variants or excess copy-number burden provided a clear alternative explanation for HSCR. Phasing of a previously defined 10 single nucleotide polymorphisms(SNPs) RET enhancer haplotype revealed inheritance of a high-risk haplotype in the proband with HSCR, whereas the proband without HSCR carried only low-risk haplotypes on both chromosomes. To place these findings in a developmental context, we analysed single-cell transcriptomic data from the developing human fetal gut and neocortex. ZEB2 and RET show overlapping expression in enteric neural crest progenitors and neuroblasts but minimal overlap in the developing neocortex, indicating that reduced RET dosage is likely to have tissue-specific effects in the enteric nervous system. Together, these results support a model in which common regulatory variation at RET modifies HSCR penetrance in the setting of ZEB2 haploinsufficiency. More broadly, our findings illustrate how whole-genome sequencing can reveal regulatory modifiers that contribute to variable expressivity in ostensibly monogenic disorders

The authors have declared no competing interest.

SCh is supported by a NIDDK R01 award DK135089, NICHD R03 award HD116004 and The Maci Whisner Research Grant from Mowat Wilson Disease Foundation. TNT is supported by NIMH R01 award MH126933 and NICHD R03 award HD116062. The funders had no role in design of the study or data interpretation

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DNA and phenotype were collected at the Sydney Childrens Hospitals Network and were approved for sequencing by Human Research Ethics Committee (SCHN HREC) and Sydney Childrens Hospitals Network Research Governance Office (SCHN RGO) under approval numbers 2019/ETH12990 and 2020/STE05398 respectively

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