Fig. 7: Identification of a CARFS351Fmutation in SCO patients.
From: CARF promotes spermatogonial self-renewal and proliferation through Wnt signaling pathway
a Sanger sequencing confirmed the CARF g.184367889 C > T mutation in the patient P2623 at the genomic DNA (gDNA) level. b Schematic representation of the domain structure of human CARF (top) and the altered amino acid identified in the mutant was colored (bottom). c H&E staining of testicular biopsies from patient P2623 and an obstructive azoospermia control. Spg, spermatogonia; Spc, spermatocyte; Spd, spermatid; Sc, Sertoli cell. (Scale bars, 15 μm). d Western blotting assays of CARF in WT and CARF−/− U2OS cells after lentivirus infection, β-actin as a loading control. Quantification of blotting intensity for indicated proteins is shown (The WT sample is set as 1.0 after normalization with β-actin). e Effects of WT CARF and CARFS351F on TOPflash reporter activity. Bar graphs represent means ± SEM. Statistical analysis was performed by two-tailed t-test, **P < 0.01. f Quantitative real-time PCR analysis of mRNA levels of Wnt target genes in WT and CARF−/− U2OS cells after lentivirus infected. Data are expressed as means ± SEM. Fold changes were compared with WT U2OS cells infected by GFP-expressing lentivirus, and were normalized to GAPDH. g H&E staining of representative testis sections from 12-month-old WT control and CARF+/− male mice (Scale bars, 200 μm). h Comparison of male fertility of 12-month-old WT control and CARF+/− mice (n = 10). Each dot in the graphs represents an individual litter. Bar graphs represent means ± SEM. Statistical analysis was performed by two-tailed t-test, ***P < 0.001. i Working model showing how CARF regulates spermatogonial self-renewal and proliferation through Wnt signaling pathway.
