Establishing the role of ZBTB20 mutations in GnRH deficiency and impaired neurogenesis in the subventricular zone: a human cohort and animal model study

Background Congenital hypogonadotropic hypogonadism (CHH) arises from defective development or dysfunction of GnRH neurons. Olfactory bulb (OB) malformations frequently accompany CHH, a condition termed Kallmann syndrome (KS). Nevertheless, approximately 50% of CHH cases remain genetically undefined. Methods Leveraging our 15-year effort in building one of Asia's largest CHH cohorts (812 unrelated patients, and 49 CHH families), we identified a pathogenic ZBTB20 mutation (p.R300C) in a four-generation CHH family with seven affected members. We generated nervous system-specific Zbtb20 conditional knockout (cKO) mice to assess phenotypic and mechanistic consequences. Molecular analyses, including transcriptional regulation assays, were performed to explore downstream pathways. Findings Four additional heterozygous ZBTB20 variants were detected in the CHH cohort. Zbtb20 cKO mice recapitulated hallmark CHH features: GnRH neuron deficiency, hypogonadism, and infertility. Notably, migration of GnRH neurons in Zbtb20 -deficient embryos was stalled along the defective terminal nerve (TN) scaffold. Furthermore, Zbtb20 deficiency impaired proliferation of neural stem cells (NSCs) in the subventricular zone (SVZ) and disrupted their migration via the rostral migratory stream (RMS) to the OB. We also identified Thbs4 as a key downstream target, as ZBTB20 transcriptionally activates Thbs4 , which is critical for SVZ NSCs migration. Interpretation Our study establishes ZBTB20 as a CHH-associated gene and demonstrates its essential role in SVZ-OB neurogenesis. Loss of ZBTB20 function leads to OB atrophy, disrupted terminal nerves, GnRH deficiency and CHH pathogenesis. Funding This project is financially supported by National Natural Science Foundation of China, and the Natural Science Foundation of Hunan Province.