Defects in GnRH Neuron Migration/Development and Hypothalamic-Pituitary Signaling Impact Clinical Variability of Kallmann Syndrome.

Kallmann syndrome (KS) is a combination of isolated hypogonadotropic hypogonadism (IHH) with olfactory dysfunction, representing a heterogeneous disorder with a broad phenotypic spectrum. The genetic background of KS has not yet been fully established. This study was conducted on 46 Polish KS subjects (41 males, 5 females; average age: 29 years old). The studied KS patients were screened for defects in a 38-gene panel with next-generation sequencing (NGS) technology. The analysis revealed 27 pathogenic and likely pathogenic (P/LP) variants, and 21 variants of uncertain significance (VUS). The P/LP variants were detected in 20 patients (43.5%). The prevalence of oligogenic P/LP defects in selected genes among KS patients was 26% (12/46), whereas the co-occurrence of other variants was detected in 43% (20 probands). The examined KS patients showed substantial genotypic and phenotypic variability. A marked difference in non-reproductive phenotypes, involving defects in genes responsible for GnRH neuron development/migration and genes contributing to pituitary development and signaling, was observed. A comprehensive gene panel for IHH testing enabled the detection of clinically relevant variants in the majority of KS patients, which makes targeted NGS an effective molecular tool. The significance of oligogenicity and the high incidence of alterations in selected genes should be further elucidated.

Compromised Volumetric Bone Density and Microarchitecture in Men With Congenital Hypogonadotropic Hypogonadism.

CONTEXT: Men with congenital hypogonadotropic hypogonadism (CHH) and Kallmann syndrome (KS) have both low circulating testosterone and estradiol levels. Whether bone structure is affected remains unknown. OBJECTIVE: To characterize bone geometry, volumetric density and microarchitecture in CHH/KS. METHODS: This cross-sectional study, conducted at a single French tertiary academic medical center, included 51 genotyped CHH/KS patients and 40 healthy volunteers. Among CHH/KS men, 98% had received testosterone and/or combined gonadotropins. High-resolution peripheral quantitative computed tomography (HR-pQCT), dual-energy x-ray absorptiometry (DXA), and measurement of serum bone markers were used to determine volumetric bone mineral density (vBMD) and cortical and trabecular microarchitecture. RESULTS: CHH and controls did not differ for age, body mass index, and levels of vitamin D and PTH. Despite long-term hormonal treatment (10.8 ± 6.8 years), DXA showed lower areal bone mineral density (aBMD) in CHH/KS at lumbar spine, total hip, femoral neck, and distal radius. Consistent with persistently higher serum bone markers, HR-pQCT revealed lower cortical and trabecular vBMD as well as cortical thickness at the tibia and the radius. CHH/KS men had altered trabecular microarchitecture with a predominant decrease of trabecular thickness. Moreover, CHH/KS men exhibited lower cortical bone area, whereas total and trabecular areas were higher only at the tibia. Earlier treatment onset (before age 19 years) conferred a significant advantage for trabecular bone volume/tissue volume and trabecular vBMD at the tibia. CONCLUSION: Both vBMD and bone microarchitecture remain impaired in CHH/KS men despite long-term hormonal treatment. Treatment initiation during adolescence is associated with enhanced trabecular outcomes, highlighting the importance of early diagnosis.

TCF12 haploinsufficiency causes autosomal dominant Kallmann syndrome and reveals network-level interactions between causal loci.

Dysfunction of the gonadotropin-releasing hormone (GnRH) axis causes a range of reproductive phenotypes resulting from defects in the specification, migration and/or function of GnRH neurons. To identify additional molecular components of this system, we initiated a systematic genetic interrogation of families with isolated GnRH deficiency (IGD). Here, we report 13 families (12 autosomal dominant and one autosomal recessive) with an anosmic form of IGD (Kallmann syndrome) with loss-of-function mutations in TCF12, a locus also known to cause syndromic and non-syndromic craniosynostosis. We show that loss of tcf12 in zebrafish larvae perturbs GnRH neuronal patterning with concomitant attenuation of the orthologous expression of tcf3a/b, encoding a binding partner of TCF12, and stub1, a gene that is both mutated in other syndromic forms of IGD and maps to a TCF12 affinity network. Finally, we report that restored STUB1 mRNA rescues loss of tcf12 in vivo. Our data extend the mutational landscape of IGD, highlight the genetic links between craniofacial patterning and GnRH dysfunction and begin to assemble the functional network that regulates the development of the GnRH axis.

Functional analysis of SEMA3A variants identified in Chinese patients with isolated hypogonadotropic hypogonadism.

Isolated hypogonadotropic hypogonadism (IHH) is a rare disorder characterized by impaired sexual development and infertility, caused by the deficiency of hypothalamic gonadotropin-releasing hormone neurons. IHH is named Kallmann's syndrome (KS) or normosmic IHH (nIHH) when associated with a defective or normal sense of smell. Variants in SEMA3A have been recently identified in patients with KS. In this study, we screened SEMA3A variants in a cohort of Chinese patients with IHH by whole exome sequencing. Three novel heterozygous SEMA3A variants (R197Q, R617Q and V458I) were identified in two nIHH and one KS patients, respectively. Functional studies indicated that R197Q and R617Q variants were ineffective in activating the phosphorylation of FAK (focal adhesion kinase) in GN11 cells, despite normal production and secretion in HEK293T cells. The V458I SEMA3A had defect in secretion as it was not detected in the conditioned medium from HEK293T cells. Compared with wild type SEMA3A protein, all three SEMA3A mutant proteins were ineffective in inducing the migration of GN11 cells. Our study further showed the contribution of SEMA3A loss-of-function variants to the pathogenesis of IHH.

Kallmann Syndrome Due to Heterozygous Mutation in SOX10 Coexisting With Waardenburg Syndrome Type II: Case Report and Review of Literature.

Introduction: Kallmann syndrome (KS) is idiopathic hypogonadotropic hypogonadism with olfactory loss or decline. Waardenburg syndrome type II (WS2) is a clinically and genetically heterogeneous disease, characterized by congenital sensorineural deafness and abnormal pigmentation of the iris, hair, and skin. Recently, mutations in the well-known WS pathogenic gene SOX10 have been found in some KS patients with deafness, but whether SOX10 is a co-pathogenic gene of KS and WS remains uncertain. Here, we report a rare case of KS and WS2 co-occurrence due to SOX10 mutations. Methods: Detailed histories were collected through questionnaires and physical examination. Blood samples of the patient and his family members were collected after obtaining informed consents. Suspected mutations were amplified and verified by Sanger sequencing after the next generation sequencing of related genes. The raw sequence data were compared to the known gene sequence data in publicly available sequence data bases using Burrows-Wheeler Aligner software (BWA, 0.7.12-r1039). Results: A 28-year-old male patient sought treatment for hypogonadism and the absence of secondary sexual characteristics. In addition, he showed signs of obesity, hyposmia, sensorineural hearing loss, and blue iris. Magnetic resonance imaging (MRI) of the olfactory bulb showed small bilateral olfactory bulbs and tracts and diaphragma cerebri. MRI of the pituitary gland revealed a flat pituitary gland in the sella. Laboratory examination demonstrated hypogonadotropic hypogonadism, pituitary hypothyroidism, subclinical hypothyroidism, and the presence of insulin resistance with normal blood glucose levels. Sequencing of the SOX10 gene showed a 20 bp insertion in between coding bases 1,179 and 1,180 (c.1179_1180insACTATGGCTCAGCCTTCCCC). This results in a frame-shifting mutation of the 394th amino acid serine in exon4 with the resulting the amino acid sequence of the protein predicted to be TMAQPSP PSPAPSLTTL TISPQDPIMA TRARPLASTR PSPIWGPRSG PSTRPSLTPA PQGPSPTAPH TGSSQYIRHC PGPKGGPVAT TPRPAPAPSL CALFLAHLRP GGGSGGG*. Conclusion: SOX10 plays an important role in some critical stages of neural crest cell development and SOX10 mutation may be a common pathogenic factor for both KS and WS. Therefore, SOX10 mutation analysis should be considered for KS patients with combined WS clinical manifestations, especially deafness.

Gonadotropin treatment for male partial congenital hypogonadotropic hypogonadism in Chinese patients.

Partial congenital hypogonadotropic hypogonadism (PCHH) is caused by an insufficiency in, but not a complete lack of, gonadotropin secretion. This leads to reduced testosterone production, mild testicular enlargement, and partial pubertal development. No studies have shown the productivity of spermatogenesis in patients with PCHH. We compared the outcomes of gonadotropin-induced spermatogenesis between patients with PCHH and those with complete congenital hypogonadotropic hypogonadism (CCHH). This retrospective study included 587 patients with CHH who were treated in Peking Union Medical College Hospital (Beijing, China) from January 2008 to September 2016. A total of 465 cases were excluded from data analysis for testosterone or gonadotropin-releasing hormone treatment, cryptorchidism, poor compliance, or incomplete medical data. We defined male patients with PCHH as those with a testicular volume of ≥4 ml and patients with a testicular volume of <4 ml as CCHH. A total of 122 compliant, noncryptorchid patients with PCHH or CCHH received combined human chorionic gonadotropin and human menopausal gonadotropin and were monitored for 24 months. Testicular size, serum luteinizing hormone levels, follicle-stimulating hormone levels, serum total testosterone levels, and sperm count were recorded at each visit. After gonadotropin therapy, patients with PCHH had a higher spermatogenesis rate (92.3%) than did patients with CCHH (74.7%). During 24-month combined gonadotropin treatment, the PCHH group took significantly less time to begin producing sperm compared with the CCHH group (median time: 11.7 vs 17.8 months, P < 0.05). In conclusion, after combined gonadotropin treatment, patients with PCHH have a higher spermatogenesis success rate and sperm concentrations and require shorter treatment periods for sperm production.

A Balanced Translocation in Kallmann Syndrome Implicates a Long Noncoding RNA, RMST, as a GnRH Neuronal Regulator.

CONTEXT: Kallmann syndrome (KS) is a rare, genetically heterogeneous Mendelian disorder. Structural defects in KS patients have helped define the genetic architecture of gonadotropin-releasing hormone (GnRH) neuronal development in this condition. OBJECTIVE: Examine the functional role a novel structural defect affecting a long noncoding RNA (lncRNA), RMST, found in a KS patient. DESIGN: Whole genome sequencing, induced pluripotent stem cells and derived neural crest cells (NCC) from the KS patient were contrasted with controls. SETTING: The Harvard Reproductive Sciences Center, Massachusetts General Hospital Center for Genomic Medicine, and Singapore Genome Institute. PATIENT: A KS patient with a unique translocation, t(7;12)(q22;q24). INTERVENTIONS/MAIN OUTCOME MEASURE/RESULTS: A novel translocation was detected affecting the lncRNA, RMST, on chromosome 12 in the absence of any other KS mutations. Compared with controls, the patient's induced pluripotent stem cells and NCC provided functional information regarding RMST. Whereas RMST expression increased during NCC differentiation in controls, it was substantially reduced in the KS patient's NCC coincident with abrogated NCC morphological development and abnormal expression of several "downstream" genes essential for GnRH ontogeny (SOX2, PAX3, CHD7, TUBB3, and MKRN3). Additionally, an intronic single nucleotide polymorphism in RMST was significantly implicated in a genome-wide association study associated with age of menarche. CONCLUSIONS: A novel deletion in RMST implicates the loss of function of a lncRNA as a unique cause of KS and suggests it plays a critical role in the ontogeny of GnRH neurons and puberty.

Live birth in male de novo Kallmann syndrome after cross-generational genetic sequencing.

PURPOSE: To present the first case proposing the use of preimplantation genetic testing for monogeneic disorders for Kallmann syndrome, providing comprehensive care in the genomic era of precision medicine. METHODS: Gonadotropin therapy was used for spermatogenesis, followed by in vitro fertilization by intracytoplasmic sperm injection and embryo transfer. Cross-generational targeted next-generation sequencing was then done for genes known to cause Kallmann syndrome. RESULTS: A heterozygous mutation at codon 102 of the FGFR1 gene was found in the patient, but the father was found to have the same mutation yet is unaffected by Kallmann syndrome. Since no causative mutation was found, a de novo or sporadic mutation was suspected as the cause of Kallmann syndrome in this case. CONCLUSIONS: Comprehensive care must be available for male Kallmann syndrome patients, as treatment should not stop at spermatogenesis, but continue with genetic counseling due to possible inheritance.

Cranial Pair 0: The Nervus Terminalis.

Originally discovered in elasmobranchs by Fritsh in 1878, the nervus terminalis has been found in virtually all species, including humans. After more than one-century debate on its nomenclature, it is nowadays recognized as cranial pair zero. The nerve mostly originates in the olfactory placode, although neural crest contribution has been also proposed. Developmentally, the nervus terminalis is clearly observed in human embryos; subsequently, during the fetal period loses some of its ganglion cells, and it is less recognizable in adults. Fibers originating in the nasal cavity passes into the cranium through the middle area of the cribiform plate of the ethmoid bone. Intracranially, fibers joint the telencephalon at several sites including the olfactory trigone and the primordium of the hippocampus to reach preoptic and precommissural regions. The nervus terminalis shows ganglion cells, that sometimes form clusters, normally one or two located at the base of the crista galli, the so-called ganglion of the nervus terminalis. Its function is uncertain. It has been described that its fibers facilitates migration of luteinizing hormone-releasing hormone cells to the hypothalamus thus participating in the development of the hypothalamic-gonadal axis, which alteration may provoke Kallmann's syndrome in humans. This review summarizes current knowledge on this structure, incorporating original illustrations of the nerve at different developmental stages, and focuses on its anatomical and clinical relevance. Anat Rec, 302:394-404, 2019. © 2018 Wiley Periodicals, Inc.

Altered White Matter Organization in the TUBB3 E410K Syndrome.

Seven unrelated individuals (four pediatric, three adults) with the TUBB3 E410K syndrome, harboring identical de novo heterozygous TUBB3 c.1228 G>A mutations, underwent neuropsychological testing and neuroimaging. Despite the absence of cortical malformations, they have intellectual and social disabilities. To search for potential etiologies for these deficits, we compared their brain's structural and white matter organization to 22 controls using structural and diffusion magnetic resonance imaging. Diffusion images were processed to calculate fractional anisotropy (FA) and perform tract reconstructions. Cortical parcellation-based network analysis and gyral topology-based FA analyses were performed. Major interhemispheric, projection and intrahemispheric tracts were manually segmented. Subjects had decreased corpus callosum volume and decreased network efficiency. While only pediatric subjects had diffuse decreases in FA predominantly affecting mid- and long-range tracts, only adult subjects had white matter volume loss associated with decreased cortical surface area. All subjects showed aberrant corticospinal tract trajectory and bilateral absence of the dorsal language network long segment. Furthermore, pediatric subjects had more tracts with decreased FA compared with controls than did adult subjects. These findings define a TUBB3 E410K neuroimaging endophenotype and lead to the hypothesis that the age-related changes are due to microscopic intrahemispheric misguided axons that are pruned during maturation.

Sema3a plays a role in the pathogenesis of CHARGE syndrome.

CHARGE syndrome is an autosomal dominant malformation disorder caused by heterozygous loss of function mutations in the chromatin remodeler CHD7. Chd7 regulates the expression of Sema3a, which also contributes to the pathogenesis of Kallmann syndrome, a heterogeneous condition with the typical features hypogonadotropic hypogonadism and an impaired sense of smell. Both features are common in CHARGE syndrome suggesting that SEMA3A may provide a genetic link between these syndromes. Indeed, we find evidence that SEMA3A plays a role in the pathogenesis of CHARGE syndrome. First, Chd7 is enriched at the Sema3a promotor in neural crest cells and loss of function of Chd7 inhibits Sema3a expression. Second, using a Xenopus CHARGE model, we show that human SEMA3A rescues Chd7 loss of function. Third, to elucidate if SEMA3A mutations in addition to CHD7 mutations also contribute to the severity of the CHARGE phenotype, we screened 31 CHD7-positive patients and identified one patient with a heterozygous non-synonymous SEMA3A variant, c.2002A>G (p.I668V). By analyzing protein expression and processing, we did not observe any differences of the p.I668V variant compared with wild-type SEMA3A, while a pathogenic SEMA3A variant p.R66W recently described in a patient with Kallmann syndrome did affect protein secretion. Furthermore, the p.I668V variant, but not the pathogenic p.R66W variant, rescues Chd7 loss of function in Xenopus, indicating that the p.I668V variant is likely benign. Thus, SEMA3A is part of an epigenetic loop that plays a role in the pathogenesis of CHARGE syndrome, however, it seems not to act as a common direct modifier.

Clinical and genetic features of 64 young male paediatric patients with congenital hypogonadotropic hypogonadism.

CONTEXT: The diagnosis of congenital hypogonadotropic hypogonadism (CHH) in prepuberty has always been challenging. Here, we aimed at studying the clinical and genetic features of paediatric CHH, especially the phenotype of hypospadias and dual defects (patients showing hypothalamic and/or pituitary defects and testicular hypoplasia), so as to have a better understanding of CHH. DESIGN: The clinical and genetic features of patients with CHH were analysed, and the relationships between hypospadias, dual defects and genetics were investigated. PATIENTS: Patients who visited Beijing Children's Hospital and were positively diagnosed with CHH. MEASUREMENTS: The collected data included sex hormones, MRI of the olfactory bulb, human chorionic gonadotrophin (hCG) test and genetic testing. We analysed clinical features and genetic results, especially hypospadias and dual defects, and compared the stimulated testosterone (T) levels in patients with and without cryptorchidism. RESULTS: Sixty-four patients were positively diagnosed, and forty-seven (73.4%) had Kallmann syndrome (KS). Four patients (6.3%) had hypospadias, including 2 KS. Micropenis combined with cryptorchidism was the most common phenotype (39%). Approximately two-third of patients showed a poor response to hCG; 15 cases were diagnosed with dual defects, and there were no significant differences between those with and without cryptorchidism. Twenty-six cases (51%) of 51 patients were identified as having classical HH mutations, affecting 10 different genes, with oligogenic mutations in 5 cases (9.8%). The most common mutations were in PROKR2 (17.6%), FGFR1 (13.7%) and CHD7 (7.8%). The frequency of PROKR2 mutations was higher in dual HH when compared to other HH cases (6/15 vs 3/36, P = .021). CONCLUSIONS: Micropenis and/or cryptorchidism can serve as important signs for the diagnosis of HH in paediatrics, and the coexistence of hypospadias does not exclude the diagnosis of CHH, including KS or normosmic isolated HH (nHH). Testicular function may be impaired earlier than expected, and PROKR2 mutations need to be evaluated to identify presumed dual defects.

Sexually dimorphic distribution of Prokr2 neurons revealed by the Prokr2-Cre mouse model.

Prokineticin receptor 2 (PROKR2) is predominantly expressed in the mammalian central nervous system. Loss-of-function mutations of PROKR2 in humans are associated with Kallmann syndrome due to the disruption of gonadotropin releasing hormone neuronal migration and deficient olfactory bulb morphogenesis. PROKR2 has been also implicated in the neuroendocrine control of GnRH neurons post-migration and other physiological systems. However, the brain circuitry and mechanisms associated with these actions have been difficult to investigate mainly due to the widespread distribution of Prokr2-expressing cells, and the lack of animal models and molecular tools. Here, we describe the generation, validation and characterization of a new mouse model that expresses Cre recombinase driven by the Prokr2 promoter, using CRISPR-Cas9 technology. Cre expression was visualized using reporter genes, tdTomato and GFP, in males and females. Expression of Cre-induced reporter genes was found in brain sites previously described to express Prokr2, e.g., the paraventricular and the suprachiasmatic nuclei, and the area postrema. The Prokr2-Cre mouse model was further validated by colocalization of Cre-induced GFP and Prokr2 mRNA. No disruption of Prokr2 expression, GnRH neuronal migration or fertility was observed. Comparative analysis of Prokr2-Cre expression in male and female brains revealed a sexually dimorphic distribution confirmed by in situ hybridization. In females, higher Cre activity was found in the medial preoptic area, ventromedial nucleus of the hypothalamus, arcuate nucleus, medial amygdala and lateral parabrachial nucleus. In males, Cre was higher in the amygdalo-hippocampal area. The sexually dimorphic pattern of Prokr2 expression indicates differential roles in reproductive function and, potentially, in other physiological systems.

A dominant negative FGFR1 mutation identified in a Kallmann syndrome patient.

Kallmann syndrome (KS) is characterized by isolated hypogonadotropic hypogonadism (IHH) with anosmia. Fibroblast growth factor receptor 1 (FGFR1) is one of KS-associated genes, accounts for approximately 10% of total patients. FGFR1 mutations have also been identified in more severe craniosynostosis syndromes, and a subset of craniosynostosis syndromes-associated FGFR1 mutations show dominant negative effect. In this study, we identified a novel FGFR1 mutation (c.867G>A; p.W289X) in a KS patient. The p.W289X mutation leads premature termination, producing a truncated FGFR1 without the transmembrane and intracellular domains. Indeed, the W289X FGFR1 was secreted into culture medium. Further, W289X FGFR1 interfered with the function of wild type receptor to induce ERK1/2 phosphorylation. We therefore identified a dominant negative FGFR1 mutation in the KS patient, and this mutant FGFR1 may be used to decipher the physiological function of FGFR1.

Anti-Müllerian Hormone and Ovarian Morphology in Women With Isolated Hypogonadotropic Hypogonadism/Kallmann Syndrome: Effects of Recombinant Human FSH.

CONTEXT: Isolated hypogonadotropic hypogonadism (IHH), characterized by gonadotropin deficiency and absent puberty, is very rare in women. IHH prevents pubertal ovarian stimulation, but anti-Müllerian hormone (AMH) and antral follicle count (AFC) have not been studied. OBJECTIVES: (1) To compare, in IHH vs controls, AMH, ovarian volume (OV), and AFC. (2) To compare, in IHH, ovarian responses to recombinant human follicle-stimulating hormone (rhFSH) and rhFSH plus recombinant human luteinizing hormone (rhLH). SUBJECTS: Sixty-eight IHH women; 51 matched healthy women. METHODS: Serum LH, FSH, sex steroids, inhibin B (InhB), AMH, and OV and AFC (sonography) were compared. Ovarian response during rhFSH administration was assessed in 12 IHH women with low AMH levels and low AFC and compared with hormonal changes observed in six additional IHH women receiving rhFSH plus rhLH. RESULTS: InhB was lower in IHH than in controls. AMH levels were also significantly lower in the patients, but two-thirds had normal values. Mean OV and total, larger, and smaller AFCs were lower in IHH than in controls. Ovarian stimulation by rhFSH led to a significant increase in serum estradiol and InhB levels and in the number of larger antral follicles. AMH and smaller AFC increased early during rhFSH stimulation but then declined despite continued stimulation. rhFSH plus rhLH stimulation led to a significantly higher increase in estradiol levels but to similar changes in circulating InhB and AMH than with rhFSH alone. CONCLUSIONS: IHH women have both low AMH levels and low AFC. However, their decrease can be reversed by follicle-stimulating hormone. Serum AMH and AFC should not serve as prognostic markers of fertility in this population.

Perinatal midline astrocyte development is impaired in fibroblast growth factor 8 hypomorphic mice.

Our previous studies showed that Fgf8 mutations can cause Kallmann syndrome (KS), a form of congenital hypogonadotropic hypogonadism, in which patients do not undergo puberty and are infertile. Interestingly, some KS patients also have agenesis of the corpus callosum (ACC) suggesting that KS pathology is not limited to reproductive function. Here, we asked whether FGF8 dysfunction is the underlying cause of ACC in some KS patients. Indeed, early studies in transgenic mice with Fgf8 mutations reported the presence of failed or incomplete corpus callosum formation. Additional studies in transgenic mice showed that FGF8 function most likely prevents the prenatal elimination of glial fibrillary acidic protein (GFAP)-immunoreactive (IR) glial cells in the indusium griseum (IG) and midline zipper (MZ), two anterior-dorsal midline regions required for corpus callosum formation (i.e., between embryonic days (E) 15.5-18.5). Here, we tested the hypothesis that FGF8 function is critical for the survival of the GFAP-IR midline glial cells. First, we measured the incidence of apoptosis in the anterior-dorsal midline region in Fgf8 hypomorphic mice during embryonic corpus callosum formation. Second, we quantified the GFAP expression in the anterior-dorsal midbrain region during pre- and postnatal development, in order to study: 1) how Fgf8 hypomorphy disrupts prenatal GFAP-IR midline glial cell development, and 2) whether Fgf8 hypomorphy continues to disrupt postnatal GFAP-IR midline glial cell development. Our results indicate that perinatal FGF8 signaling is important for the timing of the onset of anterior-dorsal Gfap expression in midline glial cells suggesting that FGF8 function regulates midline GFAP-IR glial cell development, which when disrupted by Fgf8 deficiency prevents the formation of the corpus callosum. These studies provide an experimentally-based mechanistic explanation as to why corpus callosum formation may fail in KS patients with deficits in FGF signaling.

Anosmin 1 Interacts with the Prokineticin Receptor 2 In Vitro Indicating a Molecular Link Between Both Proteins in the Pathogenesis of Kallmann Syndrome.

Sexual maturation and olfactory bulb defects found in prokineticin 2 (Pk2) and prokineticin receptor 2 (Pkr2) mutant mice resembling the phenotypic characteristics of Kallmann syndrome (KS), gave rise to the question of whether these genes would have a role in KS pathogenesis. Later, mutations in both genes were identified in patients suffering from KS. The gene responsible for the Xlinked form of KS, ANOS1, encodes the ECM protein anosmin 1. Among other functions, anosmin 1 can regulate the activity of FGFR1, encoded by one of the genes involved in the autosomal transmission of KS. Therefore, it has been proposed that anosmin 1 could interact with PKR2 to modulate its activity. We present the first evidence supporting this hypothesis and report the interaction of full-length anosmin 1 with three extracellular domains of PKR2. A truncated anosmin 1 protein comprising the first three domains of the protein interacts with the second extracellular loop of PKR2, involved in PK2 binding. Finally, last three FnIII repeats of anosmin 1 also interacted with the PKR2 domains that interacted with full-length anosmin 1. Our data represent a molecular link between two of the genes involved in KS pathogenesis.