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Neuropilin-2 regulates androgen-receptor transcriptional activity in advanced prostate cancer

Abstract

Aberrant transcriptional activity of androgen receptor (AR) is one of the dominant mechanisms for developing of castration-resistant prostate cancer (CRPC). Analyzing AR-transcriptional complex related to CRPC is therefore important towards understanding the mechanism of therapy resistance. While studying its mechanism, we observed that a transmembrane protein called neuropilin-2 (NRP2) plays a contributory role in forming a novel AR-transcriptional complex containing nuclear pore proteins. Using immunogold electron microscopy, high-resolution confocal microscopy, chromatin immunoprecipitation, proteomics, and other biochemical techniques, we delineated the molecular mechanism of how a specific splice variant of NRP2 becomes sumoylated upon ligand stimulation and translocates to the inner nuclear membrane. This splice variant of NRP2 then stabilizes the complex between AR and nuclear pore proteins to promote CRPC specific gene expression. Both full-length and splice variants of AR have been identified in this specific transcriptional complex. In vitro cell line-based assays indicated that depletion of NRP2 not only destabilizes the AR-nuclear pore protein interaction but also inhibits the transcriptional activities of AR. Using an in vivo bone metastasis model, we showed that the inhibition of NRP2 led to the sensitization of CRPC cells toward established anti-AR therapies such as enzalutamide. Overall, our finding emphasize the importance of combinatorial inhibition of NRP2 and AR as an effective therapeutic strategy against treatment refractory prostate cancer.

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Fig. 1: NRP2 localization detected within the nucleus.
Fig. 2: NRP2 present in the nuclear membrane of the cell.
Fig. 3: VEGF-C-mediated retrograde transport and post-translational SUMOylation of NRP2B.
Fig. 4: NRP2 Mass-Spectrometry predicted its interactions with various nuclear-associated proteins.
Fig. 5: Analysis of AR-NRP2 regulated gene expression.
Fig. 6: AR-DNA interaction decreases following NRP2 depletion.
Fig. 7: Inhibition of nuclear transport of NRP2 increased AR-targeted therapeutic efficacy.

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Acknowledgements

The authors thank all stuff members of the Advanced Microscopy Core and Genomics Core Facility of UNMC. We also extend our sincere thanks to the UNMC Bioinformatics and Bio-statistician Core for their support to analyze the data. The raw data has been deposited in Gene Expression Omnibus with the accession number GSE205150.

Funding

This work was supported by grants for SD (1R21CA241234-01, NE-LB506, Lageschulte Fund), KD (R01CA182435, R01CA239343, DoD W81XWH2110628), MHM and LHH (DFG project number 273676790), and MHM (DFG project number 416001651). MM is funded by the Rudolf-Becker-Foundation for his professorship. The construction of the prostate cancer tissue microarray was funded by the DFG Forschergruppe-1586 SKELMET to LCH and SF.

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SD and KD has designed the project and SD performed most of the work. NSP, RI, SB, SB, SR have assisted some of the work. TM has developed the NRP2-HA-tagged plasmid. SAA did the mass spec. AD took the electron microscopic images. MI, AB, SC, SF, MW, GBB, LCH, MHM were involved in TMA development and staining of NRP2. PG help us in promoter assay. PG, KJP, SKB and MHM critically evaluated the work and time to time provide there suggestion. DLK performed and analyzed the ChIP-seq.

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Correspondence to Samikshan Dutta or Kaustubh Datta.

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Dutta, S., Polavaram, N.S., Islam, R. et al. Neuropilin-2 regulates androgen-receptor transcriptional activity in advanced prostate cancer. Oncogene 41, 3747–3760 (2022). https://doi.org/10.1038/s41388-022-02382-y

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