Dysfunctional SEMA4D Signaling Underlies Gonadotropin-releasing Hormone Neuron Deficiency In Idiopathic Hypogonadotropic Hypogonadism

Objective This study was aimed to identify and verify a novel causal gene,SEMA4D,of idiopathic hypogonadotropic hypogonadism(IHH)with a loss-of-function mutation of SEMA4D(p.Ala515Val).We also investigated the mechanism of SEMA4D modulating GnRH neuron migration via SEMA4D/ Plexin B1/ Met/ Rnd1/ RhoA/ Raf/ MAPK signaling pathway.Our results could expand the pathogenic gene spectrum of IHH and provide the theoretical basis for gene diagnosis and individualized treatment of IHH.Methods Combination of bioinformatics,in silico and in vitro analyses was used to investigate the homozygous mutation of SEMA4D(p.Ala515Val).Functional and mechanism studies were conducted to verify the signaling pathway of SEMA4D modulating GnRH neuron migration.We used CRISPR to establish Sema4D knockout mice model to research how Sema4D effects on GnRH neuron migration,development of reproductive system and quality of sperm of mice model.Results We identified three missense mutations of SEMA4D genes with a homozygous mutation(p.Ala515Val)and verified this mutation(p.Ala515Val)a loss-of-function mutation via bioinformatics,in silico and in vitro analyses.Our results indicated the pathway,SEMA4D/Plexin B1/Met/Rnd1/RhoA/Raf/MAPK,for SEMA4D modulating GnRH neuron migration.The results of in-vivo study showed reduced GnRH neuron population in hypothalamus in Sema4D knock-out mice model with normal serum testosterone,reproductive system and quality of sperm.Conclusions This study identified and verified a novel causal gene of IHH,SEMA4D,and a loss-of-function homozygous mutation of SEMA4D.We also verified the mechanism of Sema4D promoting the migration of GnRH neurons,expanded the pathogenic gene spectrum of IHH,and provided theoretical basis for gene diagnosis and personalized treatment of IHH.