Rare double sex and mab-3-related transcription factor 1 regulatory variants in severe spermatogenic failure

Abstract

The DMRT1 (doublesex and mab-3 related transcription factor 1) gene has long been linked to sex-determining pathways across vertebrates and is known to play an essential role in gonadal development and maintenance of spermatogenesis in mice. In humans, the genomic region harboring the DMRT gene cluster has been implicated in disorders of sex development and recently DMRT1 deletions were shown to be associated with non-obstructive azoospermia (NOA). In this work we have employed different methods to screen a cohort of Portuguese NOA patients for DMRT1 exonic insertions and deletions (by MLPA; n=68) and point mutations (by Sanger sequencing; n=155). We have found three novel patient-specific non-coding variants in heterozygosity that were absent from 357 geographically matched controls. One of these is a complex variant with a putative regulatory role (c.-223_-219CGAAA>T), located in the promoter region within a conserved sequence involved in Dmrt1 repression. Moreover, while DMRT1 domains are highly conserved across vertebrates and show reduced levels of diversity in human populations, 2 rare synonymous substitutions (rs376518776 and rs34946058) and 2 rare non-coding variants that potentially affect DMRT1 expression and splicing (rs144122237 and rs200423545) were overrepresented in patients when compared to 376 Portuguese controls (301 fertile and 75 normozoospermic). Overall our previous and present results suggest a role of changes in DMRT1 dosage in NOA potentially also through a process of gene misregulation, even though DMRT1 deleterious variants seem to be rare.

CRISPR/Cas9-mediated simultaneous knockout of Dmrt1 and Dmrt3 does not recapitulate the 46,XY gonadal dysgenesis observed in 9p24.3 deletion patients

“…Thus, simultaneous loss of function of DMRT1 and DMRT3 (or DMRT1, DMRT2, and DMRT3) in those patients could have more severe effect on gonadal development than single gene knockout. Accordingly, small deletions that affect only several exons of the DMRT1 gene or regulatory variants of DMRT1 are found in patients with milder gonadal dysfunction, such as male infertility [13] , [14] .…”

Sequence analysis of 37 candidate genes for male infertility: challenges in variant assessment and validating genes

“…Its haplo-insufficiency causes a block in the elongation stage of spermiogenesis in mice, leading to asynchronous spermatid maturation (Yan et al, 2004). Deletions and missense mutations in DMRT1 are associated with a wide spectrum of phenotypes, from XY gonadal dysgenesis to disorders of spermatogenesis such as cryptozoospermia, SCO and meiotic arrest (Tewes et al, 2014;Lima et al, 2015;T€ uttelmann et al, 2018). The same variant we found, c.671A>G, has already been reported in two patients with SCO (Tewes et al, 2014), whereas missense and splicing variants in KLHL10 have only been associated with oligozoospermia in humans (Yatsenko et al, 2006;Miyamoto et al, 2016).…”

Genetic Landscape of Nonobstructive Azoospermia and New Perspectives for the Clinic

“…Finally, in 2015, the Lopes' group identified three additional noncoding variants located in regulatory regions of DMRT1, by analysing a Portuguese study cohort of 155 NOA and 376 controls (75 normozoospermic and 301 with self-reported fatherhood) using a multiplex ligation probe assay and Sanger sequencing [112]. One of them, located in the promoter region, showed clear evidence of a key regulatory role in Dmrt1 repression.…”