Abstract
BackgroundThe fibronectin type 3 and ankyrin repeat domains 1 gene, Fank1, is an ancient, evolutionarily conserved gene present in vertebrates. Short-hairpin RNA (shRNA)-based knockdown transgenic mice have oligospermia caused by an increase in apoptotic germ cells. In this study, we investigated the in vivo function of Fank1.MethodsIn this study, we generated Fank1-knockout mice using the CRISPR/Cas9 system. We then investigated the phenotype and in vivo function of Fank1. Testes and epididymis tissues were analyzed by histological and immunofluorescence staining. Apoptotic cells were analyzed in terminal deoxynucleotidyl transferase dUTP nick end-labeling assays. Fertility and sperm counts were also evaluated. The GTEx database were used to assess gene expression quantitative trait loci and mRNA expression of candidate genes and genes neighboring single nucleotide polymorphisms was analyzed by quantitative RT-PCR.ResultsIn contrast to the Fank1-knockdown model, no significant changes in epididymal sperm content and the number of apoptotic cells were observed in Fank1−/− homozygotes. In addition, a different pattern of Dusp1, Klk1b21 and Klk1b27 mRNA expression was detected in Fank1-knockout testis. These results reveal differences in the molecular changes between Fank1-knockdown mice and Fank1-knockout mice and provide a basic resource for population genetics studies.
Highlights
Genetic studies are widely used for identification of susceptibility loci in human disease (Johnson & O’Donnell, 2009)
Fank1-/- mice are fertile and have normal spermatogenesis To confirm the in vivo function of Fank1, we generated Fank1 mutant mice using the CRISPR/Cas9 system and a 70-bp deletion of exon 2 (Figs. 2A–2C)
We found that Fank1 mRNA expression levels correlated negatively with the homozygous single nucleotide polymorphisms (SNP) genotypes based on comparison with the GTEx database
Summary
Genetic studies are widely used for identification of susceptibility loci in human disease (Johnson & O’Donnell, 2009). Mouse models of gene editing are indispensable for investigations of gene function in vivo. We investigated the in vivo function of Fank. Results: In contrast to the Fank1-knockdown model, no significant changes in epididymal sperm content and the number of apoptotic cells were observed in Fank1-/- homozygotes. A different pattern of Dusp, Klk1b21 and Klk1b27 mRNA expression was detected in Fank1-knockout testis. These results reveal differences in the molecular changes between Fank1-knockdown mice and Fank1-knockout mice and provide a basic resource for population genetics studies
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
