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Updated: Aug 16, 2025
In Vivo Functional Study of Disease-associated Rare Human Variants Using Drosophila
Published on: August 20, 2019
Functional assessment of DMRT1 variants and their pathogenicity for isolated male infertility
Jana Emich1, Avinash S Gaikwad1, Birgit Stallmeyer1
1Institute of Reproductive Genetics, University of Münster, Münster, Germany.
Objective:
To study the impact of Doublesex and mab-3-related transcription factor 1 (DMRT1) gene variants on the encoded protein's function and the variants' pathogenic relevance for isolated male infertility caused by azoospermia.
Design:
This study established a novel luciferase assay for DMRT1 missense variants using 2 different target promotors and validated the assay by analyzing previously published variants associated with differences in sex development.
Setting:
University genetics research institute and tertiary referral center for couples' infertility.
Patient(S):
Eleven infertile men with severely impaired spermatogenesis resulting in crypto- or azoospermia and carrying rare heterozygous missense variants in DMRT1 were identified within the Male Reproductive Genomics study.
Main Outcome Measure(S):
Luciferase assays with human DMRT1 variants to test functional effects on the CYP19A1 and Stra8 target promoters.
Result(S):
We first developed and refined luciferase assays to reliably test the functional impact of DMRT1 missense variants. Next, the assay was validated by analyzing 2 DMRT1 variants associated with differences in sex development, of which c.240G>C p.(Arg80Ser) displayed highly significant effects on both target promoters compared with the wild-type protein (-40% and +100%, respectively) and c.331A>G p.(Arg111Gly) had a significant effect on the Stra8 promoter (-76%). We then systematically characterized 11 DMRT1 variants identified in infertile men. The de novo variant c.344T>A p.(Met115Lys) showed a pronounced loss of function in both DMRT1 target promoters (-100% and -86%, respectively). Variants c.308A>G p.(Lys103Arg) and c.991G>C p.(Asp331His) showed a significant gain of function exclusively for the CYP19A1 promoter (+15% and +19%, respectively). Based on these results, 3 variants were reclassified according to clinical guidelines.
Conclusion(S):
The present study highlights the importance of functionally characterizing DMRT1 variants of uncertain clinical significance. Using luciferase assays for diagnostic purposes enables an improved causal diagnosis for isolated male infertility.
