Extracellular vesicles carry transcriptional ‘dark matter’ revealing tissue‐specific information

• Published in: Journal of extracellular vesicles Vol. 13; no. 8; pp. e12481 - n/a
• Main Authors: Dogra, Navneet, Chen, Tzu‐Yi, Gonzalez‐Kozlova, Edgar, Miceli, Rebecca, Cordon‐Cardo, Carlos, Tewari, Ashutosh K., Losic, Bojan, Stolovitzky, Gustavo
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.08.2024; Wiley
• Subjects: Annotations; biofluids; Biopsy; Cancer; Cell Line, Tumor; Clinical trials; Dark matter; Datasets; Electron microscopes; Epithelial cells; Extracellular vesicles; Extracellular Vesicles - metabolism; Gene expression; Gene Expression Regulation, Neoplastic; Genomes; Genomics; Homeostasis; Humans; liquid biopsy; Male; Organ Specificity - genetics; Phenotypes; Prokaryotes; Prostate cancer; Prostate carcinoma; Prostatectomy; Prostatic Neoplasms - genetics; Prostatic Neoplasms - metabolism; Prostatic Neoplasms - pathology; RNA sequencing; small RNA; Transcriptome; Transcriptomes; Transcriptomics; Transmission electron microscopy; RNA sequencing; biofluids; cancer; extracellular vesicles; small RNA; liquid biopsy
• ISSN: 2001-3078; 2001-3078
• DOI: 10.1002/jev2.12481

From eukaryotes to prokaryotes, all cells secrete extracellular vesicles (EVs) as part of their regular homeostasis, intercellular communication, and cargo disposal. Accumulating evidence suggests that small EVs carry functional small RNAs, potentially serving as extracellular messengers and liquid‐biopsy markers. Yet, the complete transcriptomic landscape of EV‐associated small RNAs during disease progression is poorly delineated due to critical limitations including the protocols used for sequencing, suboptimal alignment of short reads (20–50 nt), and uncharacterized genome annotations—often denoted as the ‘dark matter’ of the genome. In this study, we investigate the EV‐associated small unannotated RNAs that arise from endogenous genes and are part of the genomic ‘dark matter’, which may play a key emerging role in regulating gene expression and translational mechanisms. To address this, we created a distinct small RNAseq dataset from human prostate cancer & benign tissues, and EVs derived from blood (pre‐ & post‐prostatectomy), urine, and human prostate carcinoma epithelial cell line. We then developed an unsupervised data‐based bioinformatic pipeline that recognizes biologically relevant transcriptional signals irrespective of their genomic annotation. Using this approach, we discovered distinct EV‐RNA expression patterns emerging from the un‐annotated genomic regions (UGRs) of the transcriptomes associated with tissue‐specific phenotypes. We have named these novel EV‐associated small RNAs as ‘EV‐UGRsʼ or “EV‐dark matter”. Here, we demonstrate that EV‐UGR gene expressions are downregulated by ∼100 fold (FDR < 0.05) in the circulating serum EVs from aggressive prostate cancer subjects. Remarkably, these EV‐UGRs expression signatures were regained (upregulated) after radical prostatectomy in the same follow‐up patients. Finally, we developed a stem‐loop RT‐qPCR assay that validated prostate cancer‐specific EV‐UGRs for selective fluid‐based diagnostics. Overall, using an unsupervised data driven approach, we investigate the ‘dark matter’ of EV‐transcriptome and demonstrate that EV‐UGRs carry tissue‐specific Information that significantly alters pre‐ and post‐prostatectomy in the prostate cancer patients. Although further validation in randomized clinical trials is required, this new class of EV‐RNAs hold promise in liquid‐biopsy by avoiding highly invasive biopsy procedures in prostate cancer.