Importin 5 (<em>Ipo5</em>) is essential to spermatogonial differentiation and fertility in adult mouse testes — ASN Events
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Importin 5 (Ipo5) is essential to spermatogonial differentiation and fertility in adult mouse testes (#93)

Penny Whiley 1 2 , Julia Young 1 2 3 , Junlan (Iris) Ma 1 , Liza O'Donnell 1 2 4 , Daniel Garama 2 5 , Mark Baker 6 , Robin M Hobbs 7 , Kate L Loveland 1 2
  1. Centre for Endocrinology and Reproductive Health, Hudson Institute of Medical Research, Clayton, VIC, Australia
  2. Department of Molecular and Translational Sciences, School of Clinical Sciences, Monash University, Clayton, VIC, Australia
  3. Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC, Australia
  4. Griffith University, Southport, Queensland, Australia
  5. Centre for Cancer Research, Hudson Institute of Medical Research, Clayton, VIC, Australia
  6. Serere Medical, Newcastle, NSW, Australia
  7. Robinson Research Institute, School of Pharmacy and Biomedical Science, Adelaide University, Adelaide, SA, Australia

Importins transport proteins into the nucleus to effect gene transcription changes; they also sequester proteins to intracellular domains and mediate BMP and WNT signalling [1, 2, 3]. Dynamic IPO5 expression and subcellular localisation are functionally linked with key male germline developmental steps, as germline-specific Ipo5 deletion in fetal (>E15.5) or newborn (PND3) testes causes germ cell depletion before/during the first round of spermatogenesis [1, 4, 5]. Here we examine IPO5 function in spermatogonia to assess its importance in adult male fertility.

In vivo analyses were conducted using our inducible Ipo5xUBC-cre knockout model (Ipo5TAM-KO). Control and Ipo5TAM-KO adult males received tamoxifen for 2 days, and testes harvested 3d (day), 1w (week), 2w, 4w, and 6w post-injection (n=3-5/timepoint). Cell type-specific markers were analysed using section immunofluorescence. RNAseq was performed on 3d, 1w, and 2w post-injection testes to identify transcriptional changes that mediate later phenotypes. Finally, potential IPO5 binding partners were identified in cultured adult mouse undifferentiated spermatogonia using immunoprecipitation and mass-spectrometry (n=3).

Ipo5TAM-KO mouse testes at 1w & 2w had more CC3/9+ cells (apoptosis, p<0.01), and fewer PLZF+ (undifferentiated spermatogonia) and GILZ+ cells (spermatogonia/spermatocytes) (p<0.05). Testis weights were progressively reduced at 4w and 6w (p<0.0001), and Sertoli cell-only tubules were abundant by 6w. RNAseq identified 87 DEGs between Control and Ipo5TAM-KO samples, most exclusive to spermatogonia/spermatocytes and relating to meiotic cell cycle and DNA repair. In agreement, IPA upstream regulator analysis implicated inhibition of STRA8, necessary for meiosis [6]. Amongst the 91 IPO5 binding proteins were histone H2A.X and SALL4, essential for undifferentiated spermatogonia maintenance [7]. Functional confirmation assessing subcellular localisation following IPO5 depletion is underway.

These investigations identify IPO5 as the first importin essential to sustaining the mouse spermatogonial/SSC population. Analysis of IPO5 binding partners will delineate regulatory proteins that underpin male germ cell biology and are required for male fertility.

  1. [1] Nathaniel, B et al. Seminars in Cell and Developmental Biology (2022)
  2. [2] Baas, R et al. The Journal of Biological Chemistry (2016)
  3. [3] Goto, T et al. The Journal of Biological Chemistry, (2013)
  4. [4] Loveland, K et al. Biology of Reproduction (2006)
  5. [5] Young, J et al. Biology of Reproduction (2025)
  6. [6] Niu, C et al. J Cell Mol Med (2020)
  7. [7] Chan, A et al. Stem Cell Reports (2017)