A homozygous variant in HFM1 causes preimplantation embryo developmental arrest by disrupting zygotic genome activation.

study questionDoes a homozygous HFM1 mutation cause human embryonic developmental arrest by disrupting zygotic genome activation? SUMMARY ANSWER: A pathogenic homozygous HFM1 mutation causes aberrant mRNA splicing and produces a protein that fails to localize to the nucleus, leading to widespread transcriptional dysregulation, failure of zygotic genome activation, and consequent embryonic arrest. WHAT IS KNOWN ALREADY: HFM1 (Helicase 1) is a germ cell-specific gene that plays a pivotal role in meiotic recombination and DNA damage repair, and its mutations are linked to premature ovarian insufficiency. While HFM1 knockout mice exhibit fertility defects, the mechanism by which HFM1 mutations cause preimplantation embryonic arrest in humans, particularly its role in zygotic genome activation, remains unclear. STUDY DESIGN, SIZE, DURATION: This was a case-based experimental study conducted from June to November 2024, involving a single infertile patient carrying a homozygous HFM1 mutation and experiencing recurrent embryonic arrest. Analyses included molecular characterization of patient embryos and functional validation in a mouse model. PARTICIPANTS/MATERIALS, SETTING,

methodsThe patient was recruited from the Reproductive Medicine Centre of the Affiliated Guangdong Second Provincial General Hospital of Jinan University. Whole-exome sequencing identified a homozygous HFM1 mutation. Minigene assays, RNA-seq, immunofluorescence, and confocal imaging were used to characterize the mutation's impact on splicing, protein localization, and transcriptomic and epigenetic states. Functional rescue experiments were performed in mouse embryos. MAIN RESULTS AND THE ROLE OF CHANCE: Functional analysis confirmed that the HFM1 mutation disrupts normal mRNA splicing, leading to the production of a protein variant that is excluded from the nucleus. Transcriptomic and epigenetic profiling of arrested human embryos linked the abnormal localization of this protein to a failure in zygotic genome activation and aberrant retention of H3K27me3. The essential role of HFM1 was further verified in a mouse model, where embryonic defects induced by HFM1 knockdown were specifically rescued by wild-type HFM1 mRNA, but not by the mutant version. LIMITATIONS, REASONS FOR CAUTION: The findings are based on a single clinical case and a limited number of embryos. Further studies with larger cohort studies are needed to validate the prevalence and pathogenicity of such mutations. Further mechanistic studies are also required to fully elucidate how HFM1 regulates gene expression and epigenetic remodeling. WIDER IMPLICATIONS OF THE

findingsThis study establishes that nuclear localization of HFM1 may be essential for ZGA and early embryogenesis in humans. It provides a mechanistic link between noncoding HFM1 variants, transcriptional dysregulation, epigenetic dysregulation, and embryonic arrest, expanding the genetic understanding of female infertility and informing future diagnostic approaches. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by National Key R&D Program of China (2022YFC2702200), the National Natural Science Foundation of China (82271728), and the Key Basic and Applied Research Project of Guangdong Province (2023B1515120027). None of the authors have any competing interests. TRIAL REGISTRATION NUMBER: n/a.