Defective signaling through plexin-A1 compromises the development of the peripheral olfactory system and neuroendocrine reproductive axis in mice.

The olfacto-genital syndrome (Kallmann syndrome) associates congenital hypogonadism due to gonadotropin-releasing hormone (GnRH) deficiency and anosmia. This is a genetically heterogeneous developmental disease with various modes of transmission, including oligogenic inheritance. Previous reports have involved defective cell signaling by semaphorin-3A in the disease pathogenesis. Here, we report that the embryonic phenotype of Plxna1-/- mutant mice lacking plexin-A1 (a major receptor of class 3 semaphorins), though not fully penetrant, resembles that of Kallmann syndrome fetuses. Pathohistological analysis indeed showed a strongly abnormal development of the peripheral olfactory system and defective embryonic migration of the neuroendocrine GnRH cells to the hypothalamic brain region in some of the mutant mice, which resulted in reduced fertility in adult males. We thus screened 250 patients for the presence of mutations in PLXNA1, and identified different nonsynonymous mutations (p.V349L, p.V437L, p.R528W, p.H684Y, p.G720E, p.R740H, p.R813H, p.R840Q, p.A854T, p.R897H, p.L1464V, p.K1618T, p.C1744F), all at heterozygous state, in 15 patients. Most of these mutations are predicted to affect plexin-A1 stability or signaling activity based on predictive algorithms and a structural model of the protein. Moreover, in vitro experiments allowed us to show the existence of deleterious effects of eight mutations (including a transcript splicing defect), none of which are expected to result in a complete loss of protein synthesis, targeting, or signaling activity, though. Our findings indicate that signaling insufficiency through plexin-A1 can contribute to the pathogenesis of Kallmann syndrome, and further substantiate the oligogenic pattern of inheritance in this developmental disorder.

Meis1 coordinates a network of genes implicated in eye development and microphthalmia.

Microphthalmos is a rare congenital anomaly characterized by reduced eye size and visual deficits of variable degree. Sporadic and hereditary microphthalmos have been associated with heterozygous mutations in genes fundamental for eye development. Yet, many cases are idiopathic or await the identification of molecular causes. Here we show that haploinsufficiency of Meis1, which encodes a transcription factor with evolutionarily conserved expression in the embryonic trunk, brain and sensory organs, including the eye, causes microphthalmic traits and visual impairment in adult mice. By combining analysis of Meis1 loss-of-function and conditional Meis1 functional rescue with ChIP-seq and RNA-seq approaches we show that, in contrast to its preferential association with Hox-Pbx BSs in the trunk, Meis1 binds to Hox/Pbx-independent sites during optic cup development. In the eye primordium, Meis1 coordinates, in a dose-dependent manner, retinal proliferation and differentiation by regulating genes responsible for human microphthalmia and components of the Notch signaling pathway. In addition, Meis1 is required for eye patterning by controlling a set of eye territory-specific transcription factors, so that in Meis1(-/-) embryos boundaries among the different eye territories are shifted or blurred. We propose that Meis1 is at the core of a genetic network implicated in eye patterning/microphthalmia, and represents an additional candidate for syndromic cases of these ocular malformations.

The prevalence of CHD7 missense versus truncating mutations is higher in patients with Kallmann syndrome than in typical CHARGE patients.

CONTEXT: Mutations in CHD7, a gene previously implicated in CHARGE (coloboma, heart defect, choanal atresia, retardation of growth and/or development, genital hypoplasia, ear anomalies) syndrome, have been reported in patients presenting with Kallmann syndrome (KS) or congenital hypogonadotropic hypogonadism (CHH). Most mutations causing CHARGE syndrome result in premature stop codons and occur de novo, but the proportion of truncating vs nontruncating mutations in KS and CHH patients is still unknown. OBJECTIVE: The objective of the study was to determine the nature, prevalence, mode of transmission, and clinical spectrum of CHD7 mutations in a large series of patients. DESIGN: We studied 209 KS and 94 CHH patients. These patients had not been diagnosed with CHARGE syndrome according to the current criteria. We searched for mutations in 16 KS and CHH genes including CHD7. RESULTS: We found presumably pathogenic mutations in CHD7 in 24 KS patients but not in CHH patients. Nontruncating mutations (16 missense and a two-codon duplication) were more prevalent than truncating mutations (three nonsense, three frame shift, and a splice site), which contrasts with patients presenting with typical CHARGE syndrome. Thus, the clinical spectrum associated with CHD7 mutations may be partly explained by genotype/phenotype correlations. Eight patients also had congenital deafness and one had a cleft lip/palate, whereas six had both. For 10 patients, the presence of diverse features of the CHARGE spectrum in at least one relative argues against a de novo appearance of the missense mutation, and this was confirmed by genetic analysis in five families. CONCLUSION: Considering the large prevalence and clinical spectrum of CHD7 mutations, it will be particularly relevant to genetic counseling to search for mutations in this gene in KS patients seeking fertility treatment, especially if KS is associated with deafness and cleft lip/palate.

Loss-of-function mutations in SOX10 cause Kallmann syndrome with deafness.

Transcription factor SOX10 plays a role in the maintenance of progenitor cell multipotency, lineage specification, and cell differentiation and is a major actor in the development of the neural crest. It has been implicated in Waardenburg syndrome (WS), a rare disorder characterized by the association between pigmentation abnormalities and deafness, but SOX10 mutations cause a variable phenotype that spreads over the initial limits of the syndrome definition. On the basis of recent findings of olfactory-bulb agenesis in WS individuals, we suspected SOX10 was also involved in Kallmann syndrome (KS). KS is defined by the association between anosmia and hypogonadotropic hypogonadism due to incomplete migration of neuroendocrine gonadotropin-releasing hormone (GnRH) cells along the olfactory, vomeronasal, and terminal nerves. Mutations in any of the nine genes identified to date account for only 30% of the KS cases. KS can be either isolated or associated with a variety of other symptoms, including deafness. This study reports SOX10 loss-of-function mutations in approximately one-third of KS individuals with deafness, indicating a substantial involvement in this clinical condition. Study of SOX10-null mutant mice revealed a developmental role of SOX10 in a subpopulation of glial cells called olfactory ensheathing cells. These mice indeed showed an almost complete absence of these cells along the olfactory nerve pathway, as well as defasciculation and misrouting of the nerve fibers, impaired migration of GnRH cells, and disorganization of the olfactory nerve layer of the olfactory bulbs.

SFRPs act as negative modulators of ADAM10 to regulate retinal neurogenesis.

It is well established that retinal neurogenesis in mouse embryos requires the activation of Notch signaling, but is independent of the Wnt signaling pathway. We found that genetic inactivation of Sfrp1 and Sfrp2, two postulated Wnt antagonists, perturbs retinal neurogenesis. In retinas from Sfrp1(-/-); Sfrp2(-/-) embryos, Notch signaling was transiently upregulated because Sfrps bind ADAM10 metalloprotease and downregulate its activity, an important step in Notch activation. The proteolysis of other ADAM10 substrates, including APP, was consistently altered in Sfrp mutants, whereas pharmacological inhibition of ADAM10 partially rescued the Sfrp1(-/-); Sfrp2(-/-) retinal phenotype. Conversely, ectopic Sfrp1 expression in the Drosophila wing imaginal disc prevented the expression of Notch targets, and this was restored by the coexpression of Kuzbanian, the Drosophila ADAM10 homolog. Together, these data indicate that Sfrps inhibit the ADAM10 metalloprotease, which might have important implications in pathological events, including cancer and Alzheimer's disease.

Differential roles of Netrin-1 and its receptor DCC in inferior olivary neuron migration.

Netrin-1 was previously shown to be required for the tangential migration and survival of neurons that will form the inferior olivary nucleus (ION). Surprisingly, the compared analysis of mutant mice lacking either Netrin-1 or its major receptor DCC reveals striking phenotypic differences besides common features. Although ectopic stops of ION cell bodies occur in the same positions along the migratory stream in both mutants, the ION neurons' number is not affected by the lack of DCC whereas it is reduced in Netrin-1 mutant mice. Thus, cell death results from the absence of Netrin-1 and not from neuron mis-routing, arguing for a role of Netrin-1 as a survival factor in vivo. The secretion of Netrin-1 by the floor plate (FP) is strictly required - whereas DCC is not - to avoid ION axons' repulsion by the FP and allows them to cross it. Leading processes of neurons of other caudal precerebellar nuclei (PCN) cannot cross the FP in either mutant mouse, suggesting differential sensitivity or mechanism of action of Netrin-1 for leading processes of ION and other PCN neurons.