# Screen of RNA binding proteins in HEK 293T cells finds 68 activators of circular RNA translation

- Published: 2026-11-05 18:16 UTC
- Section: Life & Health › Biotechnology
- Tags: circRNA translation, RNA binding proteins screen, IRES trans-acting factors, translation activators, cap-independent translation, PUF domain tethering, luciferase reporter, RNA synthetic biology, circular RNA therapeutics
- URL: https://realtech.media/news/rt-69a77fa3be044f2e98a8f6a8_screen_of_rna/

**A cell-culture screen of tethered RNA binding proteins found 68 putative activators of IRES-driven circRNA translation, and a sequence-based model predicted further candidates.**

![Screen of RNA binding proteins in HEK 293T cells finds 68 activators of circular RNA translation](https://sgzwpqqelzmajkcubgkl.supabase.co/storage/v1/object/public/rt-media/2026/11/699de7bae4925d90.jpg)
*Syed Sajidul Islam, Wikimedia Commons, CC BY-SA 4.0*

Researchers reported in Nature Communications a systematic screen of RNA binding proteins (RBPs) that enhance translation of circular RNAs (circRNAs) in human cells. The screen identified 68 proteins as putative activators of noncanonical translation [1]. The work was done in HEK 293T cells, analyzed 36 hours after transfection.

CircRNAs are closed single-stranded molecules that can be translated into proteins and, according to the authors, are more stable than linear mRNA. Properly purified circRNAs also show lower immunogenicity even without base modification, which makes them of interest for RNA therapy [1]. Their translation is mainly driven by internal ribosome entry sites (IRES) and is regulated by RBPs known as IRES trans-acting factors. However, IRES-driven translation is generally inefficient, and the factors that enhance it in circRNAs have not been thoroughly characterized [1].

## Reporter system and screen

The authors built a luminescence-based reporter. The firefly luciferase coding sequence was split into two reverse-ordered fragments flanked by introns with complementary elements. Back-splicing produces a circular RNA with an intact open reading frame, so the signal reflects translation of the circRNA rather than of linear mRNA. A Renilla luciferase gene served as a transfection control [1].

Candidate RBPs were tethered to the circRNA through a modified PUF domain that binds the 8-nucleotide sequence UUGAUAUA. Two tandem binding sites sit between the start and stop codons of the reporter. Results were normalized to a control with the PUF domain fused only to a FLAG tag. The system was validated with the translation initiation factors eIF4E and eIF4G2 [1].

## Activators and prediction

The 68 identified activators take part in RNA processing, ribosome biogenesis and translation initiation [1]. According to the authors, many of the tested RBPs promoted noncanonical translation when tethered to circRNAs. Inserting their cognate binding motifs into a circRNA can recruit the corresponding endogenous factors to drive its translation [1].

The team also trained a computational model on the experimentally tested proteins to extract common sequence features and to predict additional activators among all known RBPs. Both the newly identified and the predicted proteins promoted IRES-mediated circRNA translation, in a context-dependent manner [1]. The authors suggest these proteins may act as new IRES trans-acting factors for specific IRES elements and expand the toolbox of engineered RBPs for RNA synthetic biology [1]. The data come from cultured cells.

## Sources

1. [Nature Communications](https://doi.org/10.1038/s41467-026-77614-w)

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Source: RealTech, realtech.media — https://realtech.media/news/rt-69a77fa3be044f2e98a8f6a8_screen_of_rna/