3 minute lightning oral presentation (and poster) ESA-SRB-APEG-NZSE 2022

Examining activin and BMP signaling in human testicular cancer cell lines with emphasis on the role of nucleocytoplasmic protein IPO5 in modulating their crosstalk (#105)

Karthika Radhakrishnan 1 , Michael Luu 1 , Josephine Iaria 2 3 , Jessie M Sutherland 4 5 , Eileen A McLaughlin 4 5 , Hong-Jian Zhu 2 3 , Kate L Loveland 1 6
  1. Centre for Reproductive Health, Hudson Institute of Medical Research , Melbourne, VIC, Australia
  2. Melbourne Medical School, University of Melbourne, Melbourne, VIC, Australia
  3. Department of Surgery, Royal Melbourne Hospital, Parkville, Melbourne, VIC, Australia
  4. Hunter Medical Research Institute (HMRI), New Lambton, NSW, Australia
  5. University of Newcastle, Callaghan, NSW, Australia
  6. Dept of Molecular and Translational Sciences, Monash University, Melbourne, VIC, Australia

 

The incidence of testicular germ cell tumors (TGCTs) has steadily increased to become the most common malignancy affecting men aged between 15-40 years. Seminomas and non-seminomas (embryonal carcinomas) comprise over 95% of TGCTs [1].  These are considered to arise in the testis from fetal germ cells that fail to differentiate correctly due to genetic and/or environmental factors. Although the exact mechanism is unclear, disruption of TGF-β superfamily signaling is implicated [2]. The well-characterized cell models of seminoma (TCam-2) and embryonal carcinoma (NT2/D1) were used to study the effects of TGF-β superfamily proteins activin A and BMP4 on TGCTs. Transcript changes were measured using RNA-sequencing and RT-qPCR following 6 hours of stimulation with activin A or BMP4. Major signaling pathways and biological processes affected were identified using DAVID analyses. Activin A significantly upregulated genes linked to pluripotency, cancer, TGF-β, Notch, p53, and Hippo signaling, whereas BMP4 altered TGF-β, pluripotency, Hippo and Wnt signaling components. Predicted key upstream regulators were identified using Ingenuity Pathway Analysis software and included pharmaceuticals. Reflecting the presence of both activin and BMP in the testis in vivo, we examined the potential for their crosstalk in TCam-2 cells using a dual promoter luciferase assay and documented a dose-dependent antagonism of BMP signaling by activin A. The importin protein, IPO5, levels and subcellular localization change dramatically in fetal and adult male germline cells [3], and IPO5 selectively mediates nuclear localization of the BMP-specific SMADs1/5/9  [4]. Here we show that IPO5 knockdown significantly impeded BMP4-induced transcription of BMP-regulated genes, providing evidence that IPO5 could play a crucial role in deciding which of the two arms of TGF-β signaling is more active in TGCTs. These results demonstrate how local activin A and BMP4 levels may modulate the TGCT transcriptome and could themselves be modulated to influence testicular cancer progression. 

 

 

 

 

  1. Oosterhuis, J. W., & Looijenga, L. H. (2005). Testicular germ-cell tumours in a broader perspective. Nat Rev Cancer, 5(3), 210-222
  2. Spiller, C. M., Bowles, J., & Koopman, P. (2013). Nodal/Cripto signaling in fetal male germ cell development: implications for testicular germ cell tumors. Int J Dev Biol, 57(2-4), 211-219.
  3. Nathaniel, B., Whiley, P. A. F., Miyamoto, Y., & Loveland, K. L. (2022). Importins: Diverse roles in male fertility. Semin Cell Dev Biol, 121, 82-98.
  4. Baas, R., Sijm, A., van Teeffelen, H. A., van Es, R., Vos, H. R., & Marc Timmers, H. T. (2016). Quantitative Proteomics of the SMAD (Suppressor of Mothers against Decapentaplegic) Transcription Factor Family Identifies Importin 5 as a Bone Morphogenic Protein Receptor SMAD-specific Importin. J Biol Chem, 291(46), 24121-24132.