SEMA3A, a Gene Involved in Axonal Pathfinding, Is Mutated in Patients with Kallmann Syndrome

Kallmann syndrome (KS) associates congenital hypogonadism due to gonadotropin-releasing hormone (GnRH) deficiency and anosmia. The genetics of KS involves various modes of transmission, including oligogenic inheritance. Here, we report that Nrp1 sema/sema mutant mice that lack a functional semaphorin-binding domain in neuropilin-1, an obligatory coreceptor of semaphorin-3A, have a KS–like phenotype. Pathohistological analysis of these mice indeed showed abnormal development of the peripheral olfactory system and defective embryonic migration of the neuroendocrine GnRH cells to the basal forebrain, which results in increased mortality of newborn mice and reduced fertility in adults. We thus screened 386 KS patients for the presence of mutations in SEMA3A (by Sanger sequencing of all 17 coding exons and flanking splice sites) and identified nonsynonymous mutations in 24 patients, specifically, a frameshifting small deletion (D538fsX31) and seven different missense mutations (R66W, N153S, I400V, V435I, T688A, R730Q, R733H). All the mutations were found in heterozygous state. Seven mutations resulted in impaired secretion of semaphorin-3A by transfected COS-7 cells (D538fsX31, R66W, V435I) or reduced signaling activity of the secreted protein in the GN11 cell line derived from embryonic GnRH cells (N153S, I400V, T688A, R733H), which strongly suggests that these mutations have a pathogenic effect. Notably, mutations in other KS genes had already been identified, in heterozygous state, in five of these patients. Our findings indicate that semaphorin-3A signaling insufficiency contributes to the pathogenesis of KS and further substantiate the oligogenic pattern of inheritance in this developmental disorder.
Author Summary Kallmann syndrome is a hereditary developmental disease that affects both the hormonal reproductive axis and the sense of smell. There is a developmental link between the reproductive and olfactory disorders: neuroendocrine cells producing the gonadotropin-releasing hormone that is deficient in the patients normally migrate from the nose to the forebrain along olfactory nerve fibers during embryonic life, and they fail to do so in the patients. Affected individuals usually do not undergo spontaneous puberty. Hormone replacement therapy is the treatment to initiate virilization in males or breast development in females and later to develop fertility in both sexes. This is a genetically heterogeneous disease. Mutations in any of eight causative genes identified so far have been found in approximately 30% of the affected individuals, thus indicating that other genes remain to be discovered. We report on the identification, in 6% of the KS patients, of various loss-of-function mutations in the gene coding for semaphorin-3A, a secreted protein involved in the navigation of olfactory nerve fibers during embryogenesis. The fact that many of these mutations were also detected in clinically unaffected individuals indicates that they must combine with other genetic defects to produce the disease phenotype.