Chromatin-directed proteomics-identified network of endogenous androgen receptor in prostate cancer cells

Kaisa-Mari Launonen,Tapio Visakorpi,Jeroen Krijgsveld,Gianluca Sigismondo,Hanna Laakso,Jaana M Hartikainen,Jorma J Palvimo,Marjo Malinen,Einari A Niskanen,Reijo Sironen,Ville Paakinaho

doi:10.1038/s41388-021-01887-2

Abstract

Treatment of prostate cancer confronts resistance to androgen receptor (AR)-targeted therapies. AR-associated coregulators and chromatin proteins hold a great potential for novel therapy targets. Here, we employed a powerful chromatin-directed proteomics approach termed ChIP-SICAP to uncover the composition of chromatin protein network, the chromatome, around endogenous AR in castration resistant prostate cancer (CRPC) cells. In addition to several expected AR coregulators, the chromatome contained many nuclear proteins not previously associated with the AR. In the context of androgen signaling in CRPC cells, we further investigated the role of a known AR-associated protein, a chromatin remodeler SMARCA4 and that of SIM2, a transcription factor without a previous association with AR. To understand their role in chromatin accessibility and AR target gene expression, we integrated data from ChIP-seq, RNA-seq, ATAC-seq and functional experiments. Despite the wide co-occurrence of SMARCA4 and AR on chromatin, depletion of SMARCA4 influenced chromatin accessibility and expression of a restricted set of AR target genes, especially those involved in cell morphogenetic changes in epithelial-mesenchymal transition. The depletion also inhibited the CRPC cell growth, validating SMARCA4’s functional role in CRPC cells. Although silencing of SIM2 reduced chromatin accessibility similarly, it affected the expression of a much larger group of androgen-regulated genes, including those involved in cellular responses to external stimuli and steroid hormone stimulus. The silencing also reduced proliferation of CRPC cells and tumor size in chick embryo chorioallantoic membrane assay, further emphasizing the importance of SIM2 in CRPC cells and pointing to the functional relevance of this potential prostate cancer biomarker in CRPC cells. Overall, the chromatome of AR identified in this work is an important resource for the field focusing on this important drug target.

Highlights

Androgens and androgen receptor (AR), a hormone-activated transcription factor (TF), are key factors driving the development and progression of prostate cancer (PCa)
From the AR chromatome, we characterized and validated the roles of two proteins; SMARCA4 previously linked to the AR and single-minded homolog 2 (SIM2) hypothesized to act as a novel pioneer TF of AR
Our ChIP-SICAP quantified proteins binding to AR-containing enhancers in PCa cell milieu

Summary

Introduction

Androgens and androgen receptor (AR), a hormone-activated transcription factor (TF), are key factors driving the development and progression of prostate cancer (PCa). The AR is the primary molecular target for the hormone therapy of advanced PCa [1]. Since patients can still progress from advanced PCa to lethal castration-resistant prostate cancer (CRPC) [2], new therapeutic targets and biomarkers are needed. Even though affinity purification-coupled to mass spectrometry (MS) [6, 7] has enlightened the coregulators of NRs, it has rarely been performed in conditions that represent the natural milieu of NRs. by utilizing RIME (rapid immunoprecipitation MS of endogenous proteins), Paltoglou et al [8] and Stelloo et al [9] have identified several endogenous AR-associated proteins from cross-linked chromatin of PCa cells

Methods

Results

Conclusion