Sperm motility is vital to human reproduction. Malformations of sperm flagella can cause male infertility. Men with multiple morphological abnormalities of the flagella (MMAF) have abnormal spermatozoa with absent, short, coiled, bent, and/or irregular-caliber flagella, which impair sperm motility. The known human MMAF-associated genes, such as DNAH1, only account for fewer than 45% of affected individuals. Pathogenic mechanisms in the genetically unexplained MMAF remain to be elucidated. Here, we conducted genetic analyses by using whole-exome sequencing and genome-wide comparative genomic hybridization microarrays in a multi-center cohort of 30 Han Chinese men affected by MMAF. Among them, 12 subjects could not be genetically explained by any known MMAFassociated genes. Intriguingly, we identified compound-heterozygous mutations in CFAP43 in three subjects and a homozygous frameshift mutation in CFAP44 in one subject. All of these recessive mutations were parentally inherited from heterozygous carriers but were absent in 984 individuals from three Han Chinese control populations. CFAP43 and CFAP44, encoding two cilia-and flagella-associated proteins (CFAPs), are specifically or preferentially expressed in the testis. Using CRISPR/Cas9 technology, we generated two knockout models each deficient in mouse ortholog Cfap43 or Cfap44. Notably, both Cfap43-and Cfap44-deficient male mice presented with MMAF phenotypes, whereas the corresponding female mice were fertile. Our experimental observations on human subjects and animal models strongly suggest that biallelic mutations in either CFAP43 or CFAP44 can cause sperm flagellar abnormalities and impair sperm motility. Further investigations on other CFAP-encoding genes in more genetically unexplained MMAF-affected individuals could uncover novel mechanisms underlying sperm flagellar formation.

The corpus cavernosum is the most important structure for penile erection, and its dysfunction causes many physiological and psychological problems. However, its cellular heterogeneity and signalling networks at the molecular level are poorly understood because of limited access to samples. Here, we profile 64,993 human cavernosal single-cell transcriptomes from three males with normal erection and five organic erectile dysfunction patients. Cell communication analysis reveals that cavernosal fibroblasts are central to the paracrine signalling network and regulate microenvironmental homeostasis. Combining with immunohistochemical staining, we reveal the cellular heterogeneity and describe a detailed spatial distribution map for each fibroblast, smooth muscle and endothelial subcluster in the corpus cavernosum. Furthermore, comparative analysis and related functional experiments identify candidate regulatory signalling pathways in the pathological process. Our study provides an insight into the human corpus cavernosum microenvironment and a reference for potential erectile dysfunction therapies.

MicroRNAs (miRNAs) play important roles in mammalian spermatogenesis, which is highly dependent on Sertoli cells. However, the functions and mechanisms of miRNAs in regulating human Sertoli cells remain largely unknown. Here, we report that hsa-miR-202-3p mediates the proliferation, apoptosis, and synthesis function of human Sertoli cells. miR-202-3p was upregulated in Sertoli cells of Sertoli cell-only syndrome (SCOS) patients compared with obstructive azoospermia (OA) patients with normal spermatogenesis. Overexpression of miR-202-3p induced Sertoli cell apoptosis and inhibited cell proliferation and synthesis, and the effects were opposite when miR-202-3p was knocked down. Lipoprotein receptor-related protein 6 (LRP6) and Cyclin D1 of the Wnt/β-catenin signaling pathway were identified as direct targets of miR-202-3p in Sertoli cells, which were validated by bioinformatics tools and dual-luciferase reporter assay. Differentially expressed LRP6 and Cyclin D1 between OA and SCOS Sertoli cells were also verified. LRP6 small interfering RNA (siRNA) interference not only mimicked the effects of miR-202-3p overexpression, but also antagonized the effects of miR-202-3p inhibition on Sertoli cells. Collectively, miR-202-3p controls the proliferation, apoptosis, and synthesis function of human Sertoli cells via targeting LRP6 and Cyclin D1 of the Wnt/β-catenin signaling pathway. This study thus provides a novel insight into fate determinations of human Sertoli cells and niche of human testis.