Campos, L. A. | Search bioRxiv

bioRxiv2025

Hairpin-Functionalized Gold Nanoparticles as an Adaptable Platform for Detecting MicroRNA Signatures

Early, accurate, and fast diagnosis is essential to ensure positive health outcomes, through effective treatment interventions and disease control, as well as the study of physiological changes related to gene expression. Liquid biopsies and point-of-care (PoC) detection are valuable tools to achieve this goal, allowing the study and monitoring of a patients molecular profile in a timely and simple manner. MicroRNAs (or miRNAs) have been proposed as biomarkers for detection in liquid biopsies, as they are stable in body fluids and their dysregulation is associated with many diseases. In this work, a sensor based on gold nanoparticles (AuNPs) functionalized with oligonucleotides is reported, aiming for the detection of nucleic acids, in particular miRNAs. This sensor is based on the recognition of target sequences by hairpin-shaped oligonucleotide probes, which allows the modulation of the colloidal stability of the AuNPs, producing color changes detectable with the naked eye. The system is used to detect a panel of miRNAs, demonstrating its versatility for the detection of relevant nucleic acid signatures. The sensor detects single miRNAs with good sensitivity and selectivity and, what is more, it can be used to recognize several miRNAs simultaneously at picomolar concentrations. The system was further adapted to a lateral flow assay (LFA) format, producing a visible colored line on lateral flow test strips, and coupled with isothermal amplification to reach femtomolar detection levels. Its properties make it suitable for use in the point-of-care (PoC), contributing to fast and early detection of pathological and physiological molecular profiles.

molecular biology↗

bioRxiv2024

Virus propagation linked to exceedingly rare gene-expression errors

Viruses are obligate parasites that establish extensive interactions with proteins and other biomolecules of their hosts. About 20% of protein molecules bear phenotypic mutations due to errors during gene expression. Phenotypic mutations are not inherited and are not purged/amplified by natural selection. Therefore, protein variants harboring phenotypic mutations remain at very low levels. Here, we show that proteins at exceedingly low levels may enable virus propagation. Bacteriophage T7 recruits the host thioredoxin as an essential processivity factor for its replisome. Thioredoxin constitutive expression yields 10000-20000 molecules per E. coli cell. We inserted early stop codons in the thioredoxin gene and appended to its end the sequence encoding for a photoconvertible fluorescent protein. Virus propagation was not abolished, indicating that some thioredoxin molecules were produced through mistranscription or mistranslation. Single-molecule localization microscopy detected 12{+/-}5 molecules per cell when an ochre codon was inserted. This work demonstrates that crucial virus-host biomolecular interactions may need occur only a few times to trigger virus propagation and supports that viruses may exploit the wide diversity of host and viral protein variants arising from gene-expression errors to establish such interactions. Immediate implications of this notion for the mechanisms of cross-species transmission and antibody evasion are discussed.

microbiology↗