TY  - JOUR
AU  - Corsi, Jessica
AU  - Semnani, Pouriya Sharbatian
AU  - Peroni, Daniele
AU  - Belli, Romina
AU  - Morelli, Alessia
AU  - Lassandro, Michelangelo
AU  - Sidarovich, Viktoryia
AU  - Adami, Valentina
AU  - Valentini, Chiara
AU  - Cavallerio, Paolo
AU  - Grosskreutz, Julian
AU  - Fabbiano, Fabrizio
AU  - Grossmann, Dajana
AU  - Hermann, Andreas
AU  - Tell, Gianluca
AU  - Basso, Manuela
AU  - D'Agostino, Vito G
TI  - Small molecule inhibitors of hnRNPA2B1-RNA interactions reveal a predictable sorting of RNA subsets into extracellular vesicles.
JO  - Nucleic acids research
VL  - 53
IS  - 5
SN  - 0305-1048
CY  - Oxford
PB  - Oxford Univ. Press
M1  - DZNE-2025-00441
SP  - gkaf176
PY  - 2025
AB  - Extracellular vesicles (EVs) are cell-secreted membranous particles contributing to intercellular communication. Coding and noncoding RNAs can be detected as EV cargo, and RNA-binding proteins (RBPs), such as hnRNPA2B1, have been circumstantially implicated in EV-RNA sorting mechanisms. However, the contribution of competitive RBP-RNA interactions responsible for RNA-sorting outcomes is still unclear, especially for predicting the EV-RNA content. We designed a reverse proteomic analysis exploiting the EV-RNA to identify intracellular protein binders in vitro. Using cells expressing a recombinant hnRNPA2B1 to normalize competitive interactions, we prioritized a network of heterogeneous nuclear ribonucleoproteins and purine-rich RNA sequences subsequently validated in secreted EV-RNA through short fluorescent RNA oligos. Then, we designed a GGGAG-enriched RNA probe that efficiently interacted with a full-length human hnRNPA2B1 protein. We exploited the interaction to conduct a pharmacological screening and identify inhibitors of the protein-RNA binding. Small molecules were orthogonally validated through biochemical and cell-based approaches. Selected drugs remarkably impacted secreted EV-RNAs and reduced an RNA-dependent, EV-mediated paracrine activation of NF-kB in recipient cells. These results demonstrate the relevance of post-transcriptional mechanisms for EV-RNA sorting and the possibility of predicting the EV-RNA quality for developing innovative strategies targeting discrete paracrine functions.
KW  - Humans
KW  - Extracellular Vesicles: metabolism
KW  - Heterogeneous-Nuclear Ribonucleoprotein Group A-B: metabolism
KW  - Heterogeneous-Nuclear Ribonucleoprotein Group A-B: genetics
KW  - RNA: metabolism
KW  - RNA: genetics
KW  - Protein Binding
KW  - Proteomics: methods
KW  - HEK293 Cells
KW  - NF-kappa B: metabolism
KW  - Small Molecule Libraries: pharmacology
KW  - Heterogeneous-Nuclear Ribonucleoprotein Group A-B (NLM Chemicals)
KW  - RNA (NLM Chemicals)
KW  - hnRNP A2 (NLM Chemicals)
KW  - NF-kappa B (NLM Chemicals)
KW  - Small Molecule Libraries (NLM Chemicals)
LB  - PUB:(DE-HGF)16
C6  - pmid:40103230
DO  - DOI:10.1093/nar/gkaf176
UR  - https://pub.dzne.de/record/277552
ER  -