Glycosylation des anticorps anti-PR3 dans la polyangéite granulomateuse

CHATENAY M.-PARIS 11-BU Pharma. (920192101) / SudocSudocFranceF

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

International audienceKallmann 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

Expression of the Sema3A coreceptor Nrp1 by vomeronasal/terminal nerve fibers and migrating GnRH cells in human and mouse embryos.

<p>(A) Schematic representation of the head of a mouse embryo at E14.5, showing the scaffold of vomeronasal/terminal nerve fibers (in red) along which GnRH cells (in blue) migrate from the nose to the ventral forebrain region. Several areas along this migratory path have been shown to produce Sema3A, including the frontonasal mesenchyme and the olfactory bulb region <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002896#pgen.1002896-Schwarting1" target="_blank">[21]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002896#pgen.1002896-Giger1" target="_blank">[33]</a>. Boxes indicate the locations of the sagittal sections shown in (B) and (C). Abbreviations: oe, olfactory epithelium; vno, vomeronasal organ; nm, frontonasal mesenchyme; mob, main olfactory bulb; aob, accessory olfactory bulb; vfb, ventral forebrain; 3V, third ventricle. (B) Sagittal section of the frontonasal region in an E14.5 mouse embryo. In the frontonasal mesenchyme (nm), migrating GnRH-immunoreactive cells (green) are morphologically associated with Nrp1-immunoreactive nerve fibers (red) originating in the vomeronasal organ (vno). Single plane confocal images at higher magnification (insets) show that GnRH cells are Nrp1-immunoreactive (green+red = yellow staining). (C) Sagittal section of the ventral forebrain (vfb) in an E14.5 mouse embryo. The peripherin-immunoreactive (green) fibers of the caudal branch of the vomeronasal nerve (arrows) are also Nrp1-immunoreactive (red), as shown by their yellow staining (green+red). (D) Sagittal section of the olfactory epithelium (oe) and olfactory bulb (ob) regions (left panel) and detail of the frontonasal region (right panel) in a 9 week-old human fetus. Clusters of GnRH-immunoreactive cells (green, arrowheads) are visible in the frontonasal mesenchyme (nm) and the rostral forebrain (fb). In the frontonasal region, these cells migrate in close contact with Nrp1-immunoreactive axons (red). Note that migrating GnRH cells are also Nrp1-immunoreactive, as shown by their yellow staining (green+red) in the right panel (arrows). Scale bars: 100 µm (25 µm in insets).</p

<i>SEMA3A</i> mutations identified in Kallmann syndrome patients.

<p><i>SEMA3A</i> mutations identified in Kallmann syndrome patients.</p

Defects in olfactory and vomeronasal axons, and GnRH cell migration in <i>Nrp1</i>^sema/sema mutant mice.

<p>(A) Coronal sections of the right olfactory epithelium (oe) and olfactory bulb (ob) regions (left panels), and detail of the olfactory bulb showing the olfactory nerve layer (nl) and glomerular layer (gl) (right panels) in <i>Nrp1</i>^+/+ and <i>Nrp1</i>^sema/sema newborn (P0) mice. Axons of the olfactory receptor neurons were immunostained (red) using an antibody directed against the olfactory marker protein (OMP). In the <i>Nrp1</i>^sema/sema mouse, the immunostaining is both enlarged below the olfactory bulb ventro-medial aspect (asterisks) and markedly reduced in the glomerular layer (arrowheads) compared to wild-type. (B) Sagittal sections of the rostral and ventral forebrain regions (left panels), and detail of the caudal branch of the vomeronasal nerve (right panels) in <i>Nrp1</i>^+/+ and <i>Nrp1</i>^sema/sema E14.5 mouse embryos. A crystal of the DiI lipophilic fluorescent dye has been placed in the vomeronasal organ lumen to anterogradely label vomeronasal axons. The vomeronasal nerve extends across the medial aspect of the olfactory bulb and projects both dorsally, to the accessory olfactory bulb, and caudally, to the ventral forebrain (vfb). In the mutant mouse, fibers in the caudal branch are scarce compared to wild-type. (C) Sagittal sections of the rostral and ventral forebrain regions at E14.5, immunostained for GnRH (green). Note the abnormal distribution of GnRH-immunoreactive cells in the <i>Nrp1</i>^sema/sema mouse (arrows). (D) Coronal sections of the preoptic region (upper panels) showing GnRH neuroendocrine cells (green) and their projections in the median eminence (me, arrows) (lower panels) in <i>Nrp1</i>^+/+ and <i>Nrp1</i>^sema/sema newborn (P0) mice. The immunostaining is reduced in the <i>Nrp1</i>^sema/sema mouse. (E) Quantitative analysis (mean ± s.d.) of GnRH cell distributions in <i>Nrp1</i>^+/+ and <i>Nrp1</i>^sema/sema mice at E14.5 and P0. * and ** denote statistically significant differences between genotypes in the indicated head regions (two-way ANOVA followed by Tukey's range test) with <i>p</i><0.05 and <i>p</i><0.01, respectively. Note that the total numbers of GnRH cells are not statistically different between <i>Nrp1</i>^+/+ and <i>Nrp1</i>^sema/sema mice at E14.5 or P0 (Student's t-test, <i>p</i>>0.05). Other abbreviations: cx, cerebral cortex; ovlt, organum vasculosum of lamina terminalis; 3v, third ventricle. Scale bars: 100 µm (50 µm in inset).</p