A Pilot Study On Association Between Meiosis Related Gene HFM1 Variation And Premature Ovarian Insufficiency

Objective:Premature ovarian insufficiency(POI)is a highly heterogeneous and complex disease affected about 1% women under 40 years of age.Although it remains idiopathic in most cases,the existence of familial cases indicates that it has a strong genetic origin.The documented data shows that meiotic gene HFM1 is related to POI,but the causative mechanism is not clear.In this study,we aim to explore the possible pathogenesis of HFM1 compound heterozygous mutations(c.1686-1G > C;c.2651 T > G,p.Ile884Ser)with POI,which had been identified in our previous study.Methods:A minigene assay was performed to verify the effect of mutation(c.1686-1G > C)on pre-m RNA splicing.The mutant eukaryotic expression vectors(c.2651 T > G)of HFM1 were constructed based on the wild-type plasmids by site-directed mutagenesis to detect the level of transcriptional and translational products of HFM1 as well as the protein degradation pathway in vitro.The ATPase activity and helicase activity of wild-type and mutant HFM1 protein(p.Ile 884 Ser)were tested by purified protein derived from prokaryotic expression.Results: The result of minigene assay showed that HFM1 mutation(c.1686-1G > C)leads to the skip of the whole exon 14 on m RNA,and the RT-PCR product of mutant plasmids was 44 bp shorter than the product of wild-type.The m RNA and protein expression level in HFM1 mutant type(c.2651 T > G)were significantly lower than those in wild type.Moreover,the wild-type protein was mainly degraded through autophagy-lysosome pathway in cells,while mutant-type protein was degraded by ubiquitin-proteasome system.The ATPase activity of the mutant protein was lower than wild type proein.The helicase activity of the wild type and mutant protein was similar.Conclusions:Our results indicate that the mutation of HFM1 is a possible mechanism leading to POI,which provides meaningful clues for further research.