Fig. 2

Conditional inactivation of Uhrf1 in postnatal male germ cells results in spermatogenic arrest and male infertility in mice. a Schematic representation of the targeting strategy for generating a floxed Uhrf1 allele through homologous recombination in the murine embryonic stem cells. Exons 4 will be deleted after Cre-mediated recombination. Position of the forward (For) and reverse (Rev) primers used for genotyping are shown. b Representative PCR genotyping results showing the floxed (lox) and the WT ( + ) alleles can be detected at 173 and 264 bp bands, respectively. c RT-qPCR analyses showing Uhrf1 mRNA level was nearly undetectable and Dnmt1, Dnmt3b, and Hdac1 mRNAs levels were also dramatic downregulated in Uhrf1 cKO adult testes, whereas the expression level of Dnmt3a was upregulated. Data are presented as mean ± SEM, n = 3. *P < 0.05 by Student’s t-test. Source data are provided as a source data file. d Western blot of UHRF1, DNMT1, DNMT3A, and HDAC1 expression in adult control and Uhrf1 cKO testes. GAPDH served as a loading control. e Representative immunofluorescent images showing UHRF1 was undetectable in adult Uhrf1 cKO testis. Scale bar = 50 μm. f Gross morphology of the testis and the epididymis from control and Uhrf1 cKO mice at postnatal day 56 (P56). g Testes growth curve shows the Uhrf1 cKO testes were significantly decreased from P14. Data are presented as mean ± SEM, n = 2–5. Source data are provided as a source data file. h Periodic acid-Schiff (PAS) staining showing the histology of testis and epididymis sections from control and Uhrf1 cKO mice at P60. Upper panels indicate testicular histology of control and Uhrf1 cKO mice. Large vacuoles were seen in most seminiferous tubules of the Uhrf1 cKO testes. Lower panels showing cauda epididymis was completely lacking spermatozoa at P60 of Uhrf1 cKO mice. Scale bar = 50 μm. i Transmission electron microscopy images of control and Uhrf1 cKO testis ultra-sections at P60. Uhrf1 cKO testis sections showing massive degenerated germ cells, and very few normal spermatocytes were seen, whereas control testis sections displaying full germ cells development. Sc, Sertoli cells; Sg, Spermatogonia; Spc, Spermatocytes; Rs, Round spermatids. Scale bar = 10 μm