bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.11.24.690164

A programmable mRNA platform for miRNA detection via miRNA-mRNA2 triplex-mediated ribosomal frameshifting

Chen, Y.·Zhao, W.·Chen, H.·Zhou, H.·Fan, S.·Duan, H.·Dai, Y.·Lu, R.·Li, C.·Jiang, C.·Chan, E.·Chen, G.
View original source ↗Export metadata ↓

Abstract

Programmed -1 ribosomal frameshifting (-1 PRF) is a recoding mechanism utilized by viruses to expand their coding capacity and modulate the stoichiometric ratio of -1 frame and 0 frame translation products. The stability of mRNA secondary structure at the ribosomal entry site within the frameshifting stimulating elements (FSEs) determines the frameshifting efficiency. Here, we report the development of a programmable mRNA-based platform that detects specific mature microRNA (miRNA or miR) by converting their presence into a quantifiable protein output through miRNA-triggered -1 PRF. We designed a triplex-forming mRNA (TF-mRNA) platform to selectively trap target miRNAs through the formation of major-groove mRNA-miRNA-mRNA (miR-mRNA2) triplexes. Bio-layer interferometry and fluorescence binding studies confirmed that TF-mRNA forms stable complexes with cognate miRNAs with low nanomolar affinity and prolonged dissociation rate. Critically, the formation of miR-mRNA2 triplex robustly stimulated ribosomal frameshifting in a cell-free dual-luciferase translation system, acting as a miRNA-dependent molecular switch. The generality of this TF-mRNA platform has been verified for several disease-associated purine-rich miRNAs, and it is suitable for targeting a wide range of other purine-enriched miRNAs. This programmable TF-mRNA platform establishes a foundation for developing novel diagnostic tools and synthetic biology circuits that convert the presence of miRNA into a quantifiable protein output. TOC Figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/690164v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@1d6d9f7org.highwire.dtl.DTLVardef@8008e6org.highwire.dtl.DTLVardef@1c8344borg.highwire.dtl.DTLVardef@e153ec_HPS_FORMAT_FIGEXP M_FIG C_FIG

Source connections

Explore related subjects

synthetic biologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Chen, Y., Zhao, W., Chen, H., Zhou, H., Fan, S., Duan, H., Dai, Y., Lu, R., Li, C., Jiang, C., Chan, E., Chen, G.. 2025-11-24. A programmable mRNA platform for miRNA detection via miRNA-mRNA2 triplex-mediated ribosomal frameshifting. https://doi.org/10.1101/2025.11.24.690164

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Coupled enzyme discovery, evolution and synthetic yeast chassis adaptation for microbial biopolymer valorisation

The valorisation of biological polymers requires microbial systems that can both access recalcitrant substrates and convert the resulting carbon into useful products. Although microbial genome and metagenome resources provide an expanding reservoir of candidate depolymerizing and modifying enzymes, most discovery workflows remain disconnected from enzyme optimisation and host adaptation. Here we present a coupled sequence-based enzyme discovery, enzyme evolution and synthetic yeast chassis adaptation strategy for microbial biopolymer valorisation. Focusing on laccases for the depolymerisation of lignin as a proof of concept, we combine sequence data mining for enzyme discovery, modular yeast surface display for functional screening, directed evolution for enzyme optimisation and synthetic yeast genome diversification for chassis improvement. In our study, surface display enabled functional benchmarking and recovery of improved laccase variants and synthetic-genome-enabled diversification provided a route to explore host configurations that influence display and enzyme performance. By integrating enzyme-level and chassis-level optimisation, this framework addresses a central bottleneck in converting microbial biodiversity by computational sequence repository mining into deployable biomanufacturing systems. Our results establish laccases as tractable entry points for oxidative biopolymer conversion and provide a generalizable platform for engineering yeast systems for sustainable carbon valorisation.

synthetic biology↗
bioRxiv2026

Multichromatic Dynamic Control of Multi-Membered Microbial Consortia Compositions for Chemical Production

Engineered microbial consortia offer a promising strategy for chemical production by distributing specialized functions among microbial strains, reducing metabolic burden, facilitating modular pathway optimization, reducing toxicity, and increasing strain stability. However, differences in growth rates can destabilize population composition, compromising productivity and limiting their applicability. Here, we developed a multichromatic optogenetic Toxin-Antitoxin (optogeneticTA) platform for dynamic control of Escherichia coli consortia of up to four members using blue, red, and near-infrared light and darkness. By varying light intensities or pulses, we precisely program and dynamically modulate the population composition of two-, three-, and four-membered consortia. We further developed a modular mathematical framework that captures and predicts population dynamics of these optogenetically controlled co-cultures. Applying dynamic control to a two-membered engineered consortium increased phenol production by ~69% relative to unregulated consortia. These results establish a programmable platform for stabilizing and dynamically optimizing microbial consortia, with potential applications across microbial biomanufacturing.

synthetic biology↗
bioRxiv2026

Reassessing the contribution of the histone H3 tail to KRAB-DNMT3L-mediated epigenetic silencing

Neumann et al. introduced CHARM, a compact epigenetic silencer in which a histone H3 tail fused to DNMT3L was proposed to recruit and stimulate endogenous DNMT3A, enabling durable gene repression without a fused DNMT3A catalytic domain. Here, we evaluated the contribution of the H3 tail in independent reporter and endogenous-gene contexts. In an SNRPN reporter system, a KRAB-DNMT3L-dCas9 construct lacking the H3 tail displayed silencing kinetics comparable to CRISPRcharm Kv2, and mutating the critical H3K4 residue to alanine in CRISPRcharm Kv2 did not compromise this silencing. Similarly, after transient delivery of editor mRNAs to HEK293T cells, CRISPRcharm Kv2 did not consistently outperform the corresponding H3-tail-free construct at three endogenous loci, and mutating the critical H3K4 residue to alanine in CRISPRcharm Kv2 did not compromise this activity. These observations suggest that the engineered H3 tail does not confer a general functional advantage within the KRAB-DNMT3L-dCas9 architecture under the conditions tested.

synthetic biology↗