Perturbations of genes essential for Müllerian duct and Wölffian duct development in Mayer-Rokitansky-Küster-Hauser syndrome

Abstract

Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS) is associated with congenital absence of the uterus, cervix, and the upper part of the vagina; it is a sex-limited trait. Disrupted development of the Müllerian ducts (MD)/Wölffian ducts (WD) through multifactorial mechanisms has been proposed to underlie MRKHS. In this study, exome sequencing (ES) was performed on a Chinese discovery cohort (442 affected subjects and 941 female control subjects) and a replication MRKHS cohort (150 affected subjects of mixed ethnicity from North America, South America, and Europe). Phenotypic follow-up of the female reproductive system was performed on an additional cohort of PAX8-associated congenital hypothyroidism (CH) (n = 5, Chinese). By analyzing 19 candidate genes essential for MD/WD development, we identified 12 likely gene-disrupting (LGD) variants in 7 genes: PAX8 (n = 4), BMP4 (n = 2), BMP7 (n = 2), TBX6 (n = 1), HOXA10 (n = 1), EMX2 (n = 1), and WNT9B (n = 1), while LGD variants in these genes were not detected in control samples (p = 1.27E-06). Interestingly, a sex-limited penetrance with paternal inheritance was observed in multiple families. One additional PAX8 LGD variant from the replication cohort and two missense variants from both cohorts were revealed to cause loss-of-function of the protein. From the PAX8-associated CH cohort, we identified one individual presenting a syndromic condition characterized by CH and MRKHS (CH-MRKHS). Our study demonstrates the comprehensive utilization of knowledge from developmental biology toward elucidating genetic perturbations, i.e., rare pathogenic alleles involving the same loci, contributing to human birth defects.

Keywords: MRKHS; MRKHS, Müllerian duct; Mayer-Rokitansky-Küster-Hauser syndrome; Müllerian aplasia; PAX8; Wölffian duct.

Figure 1

Study design and workflow of the study (A) Centers of recruitment and analysis for MRKHS- and CH-affected individuals. (B) Workflow and main findings of this study. Abbreviations: MRKHS, Mayer-Rokitansky-Küster-Hauser syndrome; CH, congenital hypothyroidism; LGD, likely gene-disrupting; LoF, loss-of-function; D-mis, deleterious missense.

Figure 2

Identification of variants in genes essential for Müllerian duct (MD) and Wölffian duct (WD) development (A) The elongation process of Müllerian duct/Wölffian duct is depicted. Protein located in the corresponding regions and with prioritized variants in them are also presented. Abbreviations: LGD, likely gene-disrupting; D-mis, damaging missense. (B) Pedigree and Sanger sequencing results of three families with PAX8 variants from the discovery cohort and one family from the replication cohort. (C) Luciferase assay on PAX8 missense variants from the discovery cohort and the replication cohorts. WT, wild type; R31C, a known deleterious variant, was used as the positive control; CH, congenital hypothyroidism. The data are the mean of n = 5 independent experiments. Error bars show one standard deviation (^∗p < 0.05, Dunnett’s multiple comparison test). (D) The mutational spectrum of LGD variants and damaging missense variants in PAX8.