Dissecting the tRNA Fragment tRF3E–Nucleolin Interaction: Implications in Breast Cancer

TL;DR

This study explores how a tRNA fragment called tRF3E interacts with the protein Nucleolin (NCL) to suppress breast cancer growth.

Contribution

The study reveals the molecular mechanism of tRF3E-NCL interaction and identifies a key motif for tumor suppression.

Findings

tRF3E binds to NCL through two RNA-binding domains, forming a stable complex.

A mutation in tRF3E's 19–24 motif disrupts cooperativity and antitumor function.

tRF3E reduces breast cancer cell proliferation and colony formation.

Abstract

Nucleolin (NCL), an RNA-binding protein which regulates critical cellular processes, is frequently dysregulated in human cancers, including breast cancer, making it an attractive therapeutic target. However, molecular details of the RNA-NCL interaction have not been investigated yet. A tRNA fragment named tRF3E, displaying tumor suppressor roles in breast cancer, was found to bind NCL with high affinity displacing NCL-controlled transcripts. Here, we investigated the determinants and cooperativity of tRF3E-NCL interaction by Electrophoretic Mobility Shift Assays and in silico docking analysis, using wild-type or mutated tRF3E. We found that NCL, through its RNA-binding domains (RBD1–2 and RBD3–4), binds simultaneously two tRF3E molecules, giving rise to an energetically favored complex. Instead, a mutant form of tRF3E (M19–24), in which the NCL recognition element in position 19–24 has been disrupted, contacts NCL exclusively at RBD3–4, causing the loss of cooperativity among RBDs. Importantly, when expressed in MCF7 breast cancer cells, tRF3E significantly reduced cell proliferation and colony formation, confirming its role as tumor suppressor, but tRF3E functional properties were lost when the 19–24 motif was mutated, suggesting that cooperativity among multiple domains is required for the NCL-mediated tRF3E antitumor function. This study sheds light on the dynamic of RNA-NCL interaction and lays the foundations for using tRF3E as a promising NCL-targeted biodrug candidate.

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