Fig. 1: C12orf40 variants identified in NOA patients cause meiosis arrest and male infertility by destabilizing mRNAs required for germ cell development.
From: Loss-of-function variants in human C12orf40 cause male infertility by blocking meiotic progression
a Homozygous variants M1 and M2 identified in 3 unrelated Chinese families. The variant positions are indicated by arrow heads. b Hematoxylin & eosin staining (H&E) staining of testicular biopsy sections from wild-type (WT) and C12orf40-mutant individual (T00579:II-1). The arrow heads indicate the condensed spermatids. Scale bars, 50 μm. c Immunostaining of testicular biopsy sections from WT and C12orf40-mutant individual (T00579:II-1) for PNA (haploid spermatid marker, red) with nuclei counterstained by DAPI (blue). Scale bars, 20 μm. d Western blotting of CN725425 in indicated mouse tissues. GAPDH as internal control. e Litter size of adult males after mating with WT females (n = 6 per genotype). Error bar, mean ± SEM. ***P < 0.001. f The representative image of testes and the ratios of testis/body weight from males at 2-month-old (n = 3 per genotype). Error bar, mean ± SEM. ***P < 0.001. g H&E staining of cauda epididymis sections from adult mice. Scale bars, 50 µm. h Left, H&E staining of testis sections from adult mice, with zoomed image of spermatocytes at first MMI, as indicated by arrows. Scale bars, 50 µm. Right, Quantification of MMI spermatocytes with abnormally condensed chromatin in testes. i–k Left, immunostaining of pachytene spermatocyte spreads derived from testes for SYCP1 (i), DMC1 (j), and MLH1 (k, red) with SYCP3 (green). The arrows indicate synaptic (i) or crossover formation (k) defects. The circles indicate sex chromosomes. Scale bars, 10 µm. Right, quantification of synaptic defects on autosomes (i), DMC1 foci (j), or MLH1 foci (k) in pachytene spermatocytes. ***P < 0.001. l Left, immumohistochemical analysis of p-ser10-H3 (PH3) of paraffin sections from adult testes. Right, the percentage of tubules containing metaphase I/II spermatocytes was quantified. Scale bars, 10 μm. *P < 0.05. m DEG analysis between mutant and WT spermatocytes at LZ/PD stage. n Venn diagram showing shared downregulated genes between mutant LZ and PD spermatocytes. o EMSA using CN725425 and ssRNA substrate. p Volcano plot showing differentially enriched sites by RIP-seq using CN725425-HA knock-in and WT mouse testes. Sites with enriched HA signal (red dot) were considered as RIPed sites. q Cumulative abundance of differential expression level between mutant and WT spermatocytes of mRNAs in non-RIPed (gray) and CN725425 RIPed (blue) genes. r Venn diagram showing shared genes between CN725425-RIPed mRNAs and downregulated genes in LZ/PD spermatocytes. s Venn diagram showing CN725425-regulated RNA targets in LZ/PD spermatocytes.
