Mutations in the stromal antigen 3 (STAG3) gene cause male infertility due to meiotic arrest

Abstract

STUDY QUESTION Are sequence variants in the stromal antigen 3 (STAG3) gene a cause for non-obstructive azoospermia (NOA) in infertile human males? SUMMARY ANSWER Sequence variants affecting protein function of STAG3 cause male infertility due to meiotic arrest. WHAT IS KNOWN ALREADY In both women and men, STAG3 encodes for a meiosis-specific protein that is crucial for the functionality of meiotic cohesin complexes. Sequence variants in STAG3 have been reported to cause meiotic arrest in male and female mice and premature ovarian failure in human females, but not in infertile human males so far. STUDY DESIGN, SIZE, DURATION The full coding region of STAG3 was sequenced directly in a cohort of 28 men with NOA due to meiotic arrest. In addition, a larger group of 275 infertile men that underwent whole-exome sequencing (WES) was screened for potential STAG3 sequence variants. Furthermore, meiotic spreads, immunohistochemistry, WES and population sampling probability (PSAP) have been conducted in the index case. PARTICIPANTS/MATERIALS, SETTING, METHODS This study included 28 infertile but otherwise healthy human males who underwent Sanger sequencing of the full coding region of STAG3. Additionally, WES data of 275 infertile human males with different infertility phenotypes have been screened for relevant STAG3 variants. All participants underwent karyotype analysis and azoospermia factor (AZF) screening in advance. In the index patient, segregation analysis, WES data, PSAP, lab parameters, testis histology and nuclear spreads have been added to suplort the findings. MAIN RESULTS AND THE ROLE OF CHANCE Two compound-heterozygous variants in STAG3 (c.[1262T>G];[1312C>T], p.[(Leu421Arg)];[(Arg438Ter)]) have been found to cause male infertility due to complete bilateral meiotic arrest in an otherwise healthy human male. Compound heterozygosity was confirmed by Sanger sequencing of the parents and the patient’s brother. Other variants which may affect spermatogenesis have been ruled out through analysis of the patient’s WES data and application of the PSAP pipeline. As expected from Stag3 knockout-mice meiotic spreads, germ cells did not develop further than zygotene and showed drastic chromosome aberrations. No rare variants in STAG3 were found in the 275 infertile males with other phenotypes. Our results indicate that STAG3 variants that negatively affect its protein function are a rare cause of NOA (<1% of cases). LIMITATIONS, REASONS FOR CAUTION We identified only one patient with compound-heterozygous variants in STAG3 causing NOA due to meiotic arrest. Future studies should evaluate STAG3 variants in larger cohorts to support this finding. WIDER IMPLICATIONS OF THE FINDINGS Identification of STAG3 sequence variants in infertile human males should improve genetic counselling as well as diagnostics and treatment. Especially before testicular sperm extraction (TESE) for ICSI, STAG3 variants should be ruled out to prevent unnecessary interventions with frustrating outcomes for both patients and clinicians. STUDY FUNDING/COMPETING INTEREST(S) This work was carried out within the frame of the German Research Foundation (DFG) Clinical Research Unit ‘Male Germ Cells: from Genes to Function’ (CRU326). Work in the laboratory of R.J. is supported by a grant of the European Union H2020 program GermAge. The authors declare no conflicts of interest. TRIAL REGISTRATION NUMBER Not applicable.

Biallelic mutations in M1AP are a frequent cause of meiotic arrest leading to male infertility

“…In the initial MERGE study, the WES data of 60 highly selected, azoospermic infertile patients with unexplained, complete, bilateral germ cell arrest at the spermatocyte stage were screened for rare (MAF < 0.01 according to gnomAD-database 22 ), biallelic LoF variants. Two patients were subsequently excluded from this study, because likely causal variants in other genes had been identified in parallel: patient M870 had compound heterozygous variants in STAG3, 23 and patient M1401 had a heterozygous LoF variant in SYCP2 . 24 The prioritized genes in the remaining 58 patients were analyzed with regard to the level of expression in the testes.…”

“…The first was the X-chromosomal gene TEX11 , 6, 26 which is, according to a current structured assessment, one of only a few genes with strong clinical validity for an association with NOA. 7 Another example is the autosomal gene STAG3 , which has only recently been described in publications by us and others in parallel, 23, 27 and its clinical validity is currently ‘moderate’. Most of the proteins involved in DNA recombination, including M1AP , are highly evolutionarily conserved.…”

“…These results have been reported elsewhere. 6, 23, 24 Among the remaining group of 58 men, we identified three unrelated patients with the same homozygous frameshift variant c.676dup (p.Trp226Leufs*4) in M1AP . Thus, up to 5% of men affected by male infertility, non-obstructive azoospermia and meiotic arrest (3 out of 64) carry causal mutations in M1AP , making this a comparably common reason for germ cell arrest at the spermatocyte stage like TEX11 mutations with 6% (4 out of 64) in this selected patient population.…”

“…As an example, variants in STAG3 had previously been reported to cause POI 29 and now have recently been shown to also cause NOA and meiotic arrest. 23, 27 M1AP is predominantly expressed in the adult testis (GTEx), but also reported to be expressed in the fetal mouse ovary. 8 However, the structure of ovaries in female M1ap knockout mice appeared normal and fertility was preserved.…”

Analysis of STAG3 Mutations in Chinese Non-Obstructive Azoospermia Patients with Germ Cell Maturation Arrest

“…However, it has been reported that two SNPs (rs1727130 and rs1052482) located in the 3'-UTR of the STAG3 gene were identi ed to be associated with NOA in Korean population [30]. Furthermore, homozygous or compound-heterozygous variants of the STAG3 gene have been identi ed in NOA patients from Germany, Spain, and Australia [31][32][33]. In this study, we did not identify the same variants which may be due to the small sample size and ethnic diversity.…”