Oral Presentation ESA-SRB 2023 in conjunction with ENSA

The nuclear transport factor, Importin 5 (Ipo5) is essential for maintenance of the adult spermatogonial pool (#163)

Penny A F Whiley 1 2 , Michael M C Luu 1 , Daniel Garama 2 , Mark Baker 3 , Robin M Hobbs 1 2 , Kate L Loveland 1 2
  1. Centre for Reproductive Health, Hudson Institute of Medical Research, Clayton, Vic, Australia
  2. Department of Molecular and Translational Sciences, SubFaculty of Clinical and Molecular Medicine. Monash University, Clayton, VIC, Australia
  3. Priority Research Centre for Reproductive Science, University of Newcastle, Newcastle, NSW, Australia

Importins selectively transport proteins into the nucleus to effect gene transcription changes and can also sequester proteins to specific intracellular domains [1]. Importin 5 (IPO5) has been implicated in mediating BMP and WNT signalling [2, 3]. Its dynamic expression and subcellular localisation in maturing germ cells suggests it mediates developmental switches at key points in spermatogenesis [1, 4]. Germline-specific Ipo5 deletion after E15.5 and at PND3 both lead to germ cell loss in the first round of spermatogenesis.  Its importance in adult spermatogenesis is unknown, and we hypothesised IPO5 is vital for spermatogonial survival.

 

To assess IPO5 function in spermatogonia in vivo, we developed an inducible Ipo5 knockout model (Ipo5TAM-KO). Control and Ipo5TAM-KO adult males were injected with Tamoxifen for 2 consecutive days. Testes were harvested 2-, 4-, and 6-weeks post injection (n=3-5/timepoint). Testis tissue was analysed using immunofluorescence and real-time PCR. Additionally, IPO5 binding partners were identified in cultured adult mouse undifferentiated spermatogonia using immunoprecipitation and mass-spectrometry (n=4).

 

Testis weights of Ipo5TAM-KO mice were unaffected at 2-weeks, but progressively reduced at 4- and 6-weeks post-injection, consistent with spermatogenic failure. Ipo5 deletion efficiency was variable, so tubules were classified by IPO5 expression: normal, partial or none. Absence of IPO5 in tubules was accompanied by progressive reduction of PLZF+ (undifferentiated spermatogonia) and GILZ+ cells (spermatogonia and spermatocytes) at 2- and 4-weeks, which resulted in Sertoli cell-only tubules at 6-weeks (p<0.0001). Amongst the >60 IPO5 binding partners identified were ribosome biogenesis proteins, histones, and SALL4, essential for differentiation and long-term maintenance of undifferentiated spermatogonia [5].

 

These findings demonstrate IPO5 is required for maintenance of the adult spermatogonial pool, consistent with emerging knowledge of its roles in normal/pathological states. Ongoing investigations will identify critical molecules and pathways involved in male fertility, shedding light on the intricate roles of IPO5 in spermatogonial biology.

  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] Chan, A et al. Stem Cell Reports (2017)