Deleterious variants in X-linked RHOXF1 cause male infertility with oligo- and azoospermia.

Oligozoospermia and azoospermia are two common phenotypes of male infertility characterized by massive sperm defects owing to failure of spermatogenesis. The deleterious impact of candidate variants with male infertility is to be explored. In our study, we identified three hemizygous missense variants (c.388G>A: p.V130M, c.272C>T: p.A91V, and c.467C>T: p.A156V) and one hemizygous nonsense variant (c.478C>T: p.R160X) in the Rhox homeobox family member 1 gene (RHOXF1) in four unrelated cases from a cohort of 1201 infertile Chinese men with oligo- and azoospermia using whole-exome sequencing and Sanger sequencing. RHOXF1 was absent in the testicular biopsy of one patient (c.388G>A: p.V130M) whose histological analysis showed a phenotype of Sertoli cell-only syndrome. In vitro experiments indicated that RHOXF1 mutations significantly reduced the content of RHOXF1 protein in HEK293T cells. Specifically, the p.V130M, p.A156V, and p.R160X mutants of RHOXF1 also led to increased RHOXF1 accumulation in cytoplasmic particles. Luciferase assays revealed that p.V130M and p.R160X mutants may disrupt downstream spermatogenesis by perturbing the regulation of doublesex and mab-3 related transcription factor 1 (DMRT1) promoter activity. Furthermore, ICSI treatment could be beneficial in the context of oligozoospermia caused by RHOXF1 mutations. In conclusion, our findings collectively identified mutated RHOXF1 to be a disease-causing X-linked gene in human oligo- and azoospermia.

Identification of novel homozygous asthenoteratospermia-causing ARMC2 mutations associated with multiple morphological abnormalities of the sperm flagella.

PURPOSE: To identify the genetic causes of multiple morphological abnormalities in sperm flagella (MMAF) and male infertility in patients from two unrelated Han Chinese families. METHODS: Whole-exome sequencing was conducted using blood samples from the two individuals with MMAF and male infertility. Hematoxylin and eosin staining and scanning electron microscopy were performed to evaluate sperm morphology. Ultrastructural and immunostaining analyses of the spermatozoa were performed. The HEK293T cells were used to confirm the pathogenicity of the variants. RESULTS: We identified two novel homozygous missense ARMC2 variants: c.314C > T: p.P105L and c.2227A > G: p.N743D. Both variants are absent or rare in the human population genome data and are predicted to be deleterious. In vitro experiments indicated that both ARMC2 variants caused a slightly increased protein expression. ARMC2-mutant spermatozoa showed multiple morphological abnormalities (bent, short, coiled, absent, and irregular) in the flagella. In addition, the spermatozoa of the patients revealed a frequent absence of the central pair complex and disrupted axonemal ultrastructure. CONCLUSION: We identified two novel ARMC2 variants that caused male infertility and MMAF in Han Chinese patients. These findings expand the mutational spectrum of ARMC2 and provide insights into the complex causes and pathogenesis of MMAF.