A DNA-Fueled and Catalytic Molecule Machine Lights Up Trace Under-Expressed MicroRNAs in Living Cells.

Abstract

The detection of specific intracellular microRNAs (miRNAs) in living cells can potentially provide insight into the causal mechanism of cancer metastasis and invasion. However, because of the characteristic nature of miRNAs in terms of small sizes, low abundance, and similarity among family members, it is a great challenge to monitor miRNAs in living cells, especially those with much lower expression levels. In this work, we describe the establishment of a DNA-fueled and catalytic molecule machinery in cell signal amplification approach for monitoring trace and under-expressed miRNAs in living cells. The presence of the target miRNA releases the hairpin sequences from the dsDNA (containing the fluorescence resonance energy transfer (FRET) pair-labeled and unfolded hairpin sequences)-conjugated gold nanoparticles (dsDNA-AuNPs), and the DNA fuel strands assist the recycling of the target miRNA sequences via two cascaded strand displacement reactions, leading to the operation of the molecular machine in a catalytic fashion and the release of many hairpin sequences. As a result, the liberated hairpin sequences restore the folded hairpin structures and bring the FRET pair into close proximity to generate significantly amplified signals for detecting trace miRNA targets. Besides, the dsDNA-AuNP nanoprobes have good nuclease stability and show low cytotoxicity to cells, and the application of such a molecular system for monitoring trace and under-expressed miRNAs in living cells has also been demonstrated. With the advantages of in cell signal amplification and reduced background noise, the developed method thus offers new opportunities for detecting various trace intracellular miRNA species.