Oral Presentation ESA-SRB-NZSE-CaSR 2026 in conjunction with ENSA

The role of TIMM44 in premature ovarian insufficiency (143614)

Xuebi Cai 1 , Brianna Kline 1 , Firman Idris 1 , Izaac Moran 2 , Lucas Ferreira 1 , Jocelyn Van Den Bergen 1 , Gorjana Robevska 1 , Linda Akloul 3 , Bénédicte Nouyou 4 , Eve Besnier 4 , Fernando Wijaya 2 , Qiaolin Yang 2 , Justin Chen 2 , Karla Hutt 5 , John Carroll 5 , Katie Ayers 1 6 , Andrew Sinclair 1 6 , Gary Hime 2 , Sylvie Jaillard 7 , Elena Tucker 1 6
  1. Murdoch Children's Resaerch Institute, Parkville, VICTORIA, Australia
  2. Department of Anatomy and Physiology, The University of Melbourne, Melbourne, Victoria, Australia
  3. service de Génétique clinique, Univ Rennes, CHU Rennes, Rennes, Brittany (Bretagne), France
  4. service de Cytogénétique et Biologie cellulaire, Univ Rennes, CHU Rennes, Rennes, Brittany (Bretagne), France
  5. Ovarian Biology Laboratory, Biomedicine Discovery Institute, Department of Anatomy and Developmental Biology, Monash University, Melbourne, Victoria, Australia
  6. Department of Paediatrics, University of Melbourne, Melbourne, Victoria, Australia
  7. Inserm, Irset (institut de recherche en santé, environnement et travail) – UMR_S 1085, (service de Cytogénétique et Biologie cellulaire), Univ Rennes, CHU Rennes, Rennes, Brittany (Bretagne), France

Background: Premature ovarian insufficiency (POI) is characterized by the loss of normal ovarian function before the age of 40 years. Increasing evidence implicates mitochondrial dysfunction in ovarian aging and follicular depletion, suggesting that defects in mitochondrial proteins contribute to POI. Translocase of the Inner Mitochondrial Membrane 44 (TIMM44) is a key component of the mitochondrial protein import machinery; however, its role in POI remains unexplored.

Methods: Long read Oxford Nanopore sequencing was performed in a patient diagnosed with POI at 15 years of age who presented with secondary amenorrhea, atrophic ovaries and markedly elevated follicle-stimulating hormone (FSH=102 IU/L). A drosophila melanogaster knockdown (KD) model targeting the orthologue of human TIMM44 was generated using RNA interference to investigate its role in ovarian function. In parallel, A TIMM44-knockout (KO) HeLa cell line was generated using CRISPR/Cas9 genome editing to assess the functional consequences of the identified variants. Cell viability and mitochondrial function were evaluated in both wild-type (WT) and TIMM44-KO cells, followed by rescue experiments using lentiviral overexpression of WT TIMM44 or mutant TIMM44 constructs.

Results: Compound heterozygous TIMM44 variants, c.829C>G, (p.Arg277Gly) and c.541del, p.(Gln181Argfs*6)), were identified. In Drosophila, knockdown of the TIMM44 orthologue in either germ cells or somatic cells resulted in significant ovarian structural abnormalities and reduced female fertility. Furthermore, the TIMM44 orthologue was required for normal testis integrity and male fertility, indicating a conserved role in reproductive function. In human cells, TIMM44 deficiency caused mitochondrial dysfunction, evidenced by significantly reduced expression of oxidative phosphorylation complexes I, IV, and V.

Conclusion: Our findings identify TIMM44 as a novel candidate gene associated with POI and demonstrate its essential role in mitochondrial function and fertility. Functional studies in human cells and Drosophila support a conserved role for TIMM44 in reproductive biology and provide new insights into the mitochondrial mechanisms underlying female infertility.