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ABSTRACT: Müllerian inhibiting substance (MIS), a secreted glycoprotein in the transforming growth factor—beta family of growth factors, mediates regression of the Müllerian ducts during embryonic sex differentiation in males. In persistent Müllerian duct syndrome (PMDS), rather than undergoing involution, the Müllerian ducts persist in males, giving rise to the uterus, fallopian tubes, and upper vagina. Genetic defects in MIS or its receptor (MISRII) have been identified in patients with PMDS. The phenotype in the canine model of PMDS derived from the miniature schnauzer breed is strikingly similar to that of human patients. In this model, PMDS is inherited as a sex-limited autosomal recessive trait. Previous studies indicated that a defect in the MIS receptor or its downstream signaling pathway was likely to be causative of the canine syndrome. In this study, the canine PMDS phenotype and clinical sequelae are described in detail. Affected and unaffected members of this pedigree are genotyped, identifying a single base pair substitution in MISRII that introduces a stop codon in exon 3. The homozygous mutation terminates translation at 80 amino acids, eliminating much of the extracellular domain and the entire transmembrane and intracellular signaling domains. Findings in this model could enable insights to be garnered from correlation of detailed clinical descriptions with molecular defects, which are not otherwise possible in the human syndrome.
During sexual differentiation of male embryos, Müllerian inhibiting substance (MIS), also called anti-Müllerian hormone (AMH), plays an important role in mediating regression of the Müllerian ducts, the embryologic precursors of the uterus, fallopian tubes, and upper vagina (Jost, 1953). MIS is a secreted glycoprotein in the transforming growth factor—beta (TGF-beta) family of growth factors, which was identified initially for its fetal role in Müllerian duct regression but has been found to have additional postnatal roles in gonad development (Racine et al, 1998; Durlinger et al, 1999; Lee et al, 1999; Wu et al, 2005). Like other members of the TGF-beta family, MIS signals through a receptor complex comprised of type I and II serine-threonine kinase receptors. MIS binds to its type II receptor (MISRII), which has a cysteine-rich extracellular domain that confers ligand specificity and an intracellular kinase domain (Baarends et al, 1994; di Clemente et al, 1994; Teixeira and Donahoe, 1996). Upon ligand binding, MISRII recruits and phosphorylates a shared, ligand-independent type I receptor, which initiates downstream activation of intracellular mediators such as Smad1, 5, and 8 (Gouedard et al, 2000; Visser et al, 2001; Jamin et al, 2002) other signaling pathways, or both, such as beta-catenin (Allard et al, 2000) and nuclear factor kB (Segev et al, 2002). The ability of MIS to induce Müllerian duct regression is thought to be mediated via a paracrine mechanism involving mesothelial-epithelial interactions. Although MISRII is expressed in the mesenchymal cells, it is the adjacent epithelial cells that undergo apoptotic cell death during Müllerian duct regression (Allard et al, 2000; Xavier and Allard, 2003). The exact signaling pathway has not been delineated; nevertheless, the expression of biologically active MIS and its receptors during a critical embryonic window is essential for normal male-specific internal reproductive tract development.
A defect in MIS signaling causes retained rudimentary Müllerian structures or an infantile uterus and Fallopian tubes, a condition found in humans (Brook et al, 1973; Sloan and Walsh, 1976) as well as a number of other species such as dogs (Brown et al, 1976), cattle (Jost, 1965), goats (Haibel and Rojko, 1990), and cats (Schulman and Levine, 1989). In infants and children, this is typically identified at the time of surgery for cryptorchidism or inguinal hernia and is termed persistent Müllerian duct syndrome (PMDS; Brook et al, 1973; Sloan and Walsh, 1976; Josso et al, 1983). The testes can be bilaterally undescended, or one can descend and carry the contralateral testis into the same scrotum, a condition called transverse testicular ectopia.
In patients with PMDS, the mode of inheritance is primarily autosomal recessive. Molecular studies have identified genetic defects in either MIS or MISRII in over 80% of patients with PMDS (Guerrier et al, 1989; Imbeaud et al, 1994, 1995; Messika-Zeitoun et al, 2001; Belville et al, 2004). Those patients with MIS gene defects have unmeasurable or low serum concentrations of MIS and have been found to have a number of different mutations spanning the gene. In contrast, mutations of MISRII are more conserved, with a common 27-bp deletion in exon 10 in the serine-threonine kinase domain, accounting for 25% of known receptor defects (Imbeaud et al, 1996). The MISRII mutations identified thus far either affect ligand binding or abrogate kinase activity of the receptor with no mutations of the transmembrane region (exon 4) identified as yet.
Canine PMDS has been reported as an inherited disorder in 2 breeds: the miniature schnauzer in the United States (Brown et al, 1976; Marshall et al, 1982; Meyers-Wallen et al, 1989) and the basset hound in Europe (Nickel et al, 1992). The causative genetic defect has not been identified in either breed. The canine model derived from the PMDS miniature schnauzer has a phenotype that is strikingly similar to that of human PMDS (Meyers-Wallen et al, 1989). In the PMDS model, the expression of MIS mRNA and protein were no different in testes of PMDS embryos than those of normal embryos during the critical period for Müllerian duct regression (Meyers-Wallen et al, 1991, 1993). Furthermore, in studies for which a semi-quantitative urogenital ridge organ culture bioassay for MIS activity was used (Donahoe et al, 1976, 1977a,b,c; Meyers-Wallen et al, 1989), testes from affected neonates and embryos were found to have comparable MIS bioactivity during Müllerian duct regression as age-matched testes of normal dogs, confirming that the MIS was biologically functional. These findings indicated that target organ insensitivity caused by a mutation in either MISRII or a downstream gene in its signaling pathway was likely to be causative (Meyers-Wallen et al, 1993). In this study, we delineate the phenotype and clinical sequelae of this canine PMDS model in greater detail and identify the causative molecular defect in the pedigree with sex-limited autosomal recessive PMDS.