A novel mechanism of SRRM4 in promoting neuroendocrine prostate cancer development via a pluripotency gene network

Abstract

Background Prostate adenocarcinoma (AdPC) cells can undergo lineage switching to neuroendocrine cells and develop into therapy-resistant neuroendocrine prostate cancer (NEPC). While genomic/epigenetic alterations are shown to induce neuroendocrine differentiation via an intermediate stem-like state, RNA splicing factor SRRM4 can transform AdPC cells into NEPC xenografts through a direct neuroendocrine transdifferentiation mechanism. Whether SRRM4 can also regulate a stem-cell gene network for NEPC development remains unclear. Methods Multiple AdPC cell models were transduced by lentiviral vectors encoding SRRM4. SRRM4-mediated RNA splicing and neuroendocrine differentiation of cells and xenografts were determined by qPCR, immunoblotting, and immunohistochemistry. Cell morphology, proliferation, and colony formation rates were also studied. SRRM4 transcriptome in the DU145 cell model was profiled by AmpliSeq and analyzed by gene enrichment studies. Findings SRRM4 induces an overall NEPC-specific RNA splicing program in multiple cell models but creates heterogeneous transcriptomes. SRRM4-transduced DU145 cells present the most dramatic neuronal morphological changes, accelerated cell proliferation, and enhanced resistance to apoptosis. The derived xenografts show classic phenotypes similar to clinical NEPC. Whole transcriptome analyses further reveal that SRRM4 induces a pluripotency gene network consisting of the stem-cell differentiation gene, SOX2. While SRRM4 overexpression enhances SOX2 expression in both time- and dose-dependent manners in DU145 cells, RNA depletion of SOX2 compromises SRRM4-mediated stimulation of pluripotency genes. More importantly, this SRRM4-SOX2 axis is present in a subset of NEPC patient cohorts, patient-derived xenografts, and clinically relevant transgenic mouse models. Interpretation We report a novel mechanism by which SRRM4 drives NEPC progression via a pluripotency gene network. Fund Canadian Institutes of Health Research, National Nature Science Foundation of China, and China Scholar Council.

Alternative RNA splicing of the GIT1 gene is associated with neuroendocrine prostate cancer

“…Interestingly, this SRRM4‐directed RNA splicing profile shares a similar pattern to the diverse alternative splicing patterns seen in the neural system during development, where SRRM4‐spliced target genes have recognized functions that are crucial for neural programs early in development . Furthermore, our studies have reported that this reprogrammed RNA splicing signature is unique to NEPC patient tumors, PDX models, and cell models, indicating a clear variance in the phenotype of NEPC and AdPC tumors . The SRRM4‐driven t‐NEPC‐specific reprogramming of the transcriptome modifies anti‐apoptotic factors (eg, Bif‐1), epigenetic modifiers (eg, MEAF6‐1), and transcriptional regulators (eg, REST, FOXA1) that are important for regulating cell survival, proliferation and tumorigenesis, and neural differentiation, respectively .…”

“…The BPH‐1 cell line was provided by Dr Simon Hayward (Vanderbilt University, TN, USA). The LnNE and DuNE cell models, along with their control cells, were previously established by our group . All cell‐culturing conditions have been previously reported …”

“…The CRPC TMA containing 64 tissue cores from 32 patients that had undergone hormonal therapy, chemotherapy, or radiotherapy was obtained from the tissue bank at VPC. Histopathology of typical AdPC (n = 44), AdNC (atypical AdPC with ≥10% NE cells; n = 6), and SCNC (small‐cell NEPC with NE cells only; n = 6) has previously been reported and characterized …”

“…RNA in situ hybridization probes targeting GIT1‐A (944‐985 bp; ) and GIT1‐C (941‐983 bp; ) were designed by Advanced Cell Diagnostics (Hayward, CA, USA). Optimization of RISH probes and RISH probes targeting SRRM4 has previously been characterized . RISH results are available in Table .…”

“…The SRRM4‐driven t‐NEPC‐specific reprogramming of the transcriptome modifies anti‐apoptotic factors (eg, Bif‐1), epigenetic modifiers (eg, MEAF6‐1), and transcriptional regulators (eg, REST, FOXA1) that are important for regulating cell survival, proliferation and tumorigenesis, and neural differentiation, respectively . Together, these studies propose a model in which SRRM4 drives t‐NEPC development through alternative splicing of downstream genes …”