Live-Cell Imaging of MiRNAs with the Combination of CHA and HCR Techniques

Overview

Journal Methods Mol Biol

Specialty Molecular Biology

Date 2024 Nov 13

PMID 39535649

Authors

Zai-Sheng Wu

Congcong Li

Affiliations

Soon will be listed here.

Abstract

The abnormal expression of miRNA-21 is closely related to many cancers, and its sensitive detection plays an important role in the diagnosis and treatment of cancers. In the report, we designed a non-enzymatic amplification sensing system based on the catalytic hairpin assembly (CHA) and palindrome-based hybridization chain reaction (PHCR) technique for the detection and imaging of miRNA in living cells. Based on PHCR technology, two ingeniously designed palindrome-contained fluorescently labeled hairpin probes are sequentially opened upon the stimulation of short oligonucleotide triggers, promoting the autonomous assembly of cross-linked network-like structures (CNSs) and generating fluorescence resonance energy transfer (FRET) signal. Moreover, the PHCR technique can be combined with CHA technology to sufficiently improve amplification efficiency and signal output. By utilizing the CHA-PHCR sensing system, one miRNA-21 activates many triggers through CHA process and in turn initiates the assembly of CNSs by PHCR reaction, efficiently amplifying the FRET signal output. As such, the sensing system can detect target miRNAs down to 10 pm with high detection specificity. Moreover, the CNS exhibits the enhanced intracellular stability, enabling the living cell imaging of miRNA-21. The CHA-PHCR sensing system can also screen the expression level of miRNA-21 in different living cells and differentiate cancer cells from healthy cells, which is consistent with the gold-standard PCR method.

References

Cheloufi S, Dos Santos C, Chong M, Hannon G . A dicer-independent miRNA biogenesis pathway that requires Ago catalysis. Nature. 2010; 465(7298):584-9. PMC: 2995450. DOI: 10.1038/nature09092. View

Ong S, Lee W, Kodo K, Wu J . MicroRNA-mediated regulation of differentiation and trans-differentiation in stem cells. Adv Drug Deliv Rev. 2015; 88:3-15. PMC: 4546706. DOI: 10.1016/j.addr.2015.04.004. View

Shimono Y, Zabala M, Cho R, Lobo N, Dalerba P, Qian D . Downregulation of miRNA-200c links breast cancer stem cells with normal stem cells. Cell. 2009; 138(3):592-603. PMC: 2731699. DOI: 10.1016/j.cell.2009.07.011. View

de Leeuw D, Verhagen H, Denkers F, Kavelaars F, Valk P, Schuurhuis G . MicroRNA-551b is highly expressed in hematopoietic stem cells and a biomarker for relapse and poor prognosis in acute myeloid leukemia. Leukemia. 2015; 30(3):742-6. DOI: 10.1038/leu.2015.160. View

Adams B, Anastasiadou E, Esteller M, He L, Slack F . The Inescapable Influence of Noncoding RNAs in Cancer. Cancer Res. 2015; 75(24):5206-10. PMC: 4823153. DOI: 10.1158/0008-5472.CAN-15-1989. View

Zhang J, Xiao X, Liu J . The role of circulating miRNAs in multiple myeloma. Sci China Life Sci. 2015; 58(12):1262-9. DOI: 10.1007/s11427-015-4969-2. View

Dirks R, Pierce N . Triggered amplification by hybridization chain reaction. Proc Natl Acad Sci U S A. 2004; 101(43):15275-8. PMC: 524468. DOI: 10.1073/pnas.0407024101. View

Xuan F, Hsing I . Triggering hairpin-free chain-branching growth of fluorescent DNA dendrimers for nonlinear hybridization chain reaction. J Am Chem Soc. 2014; 136(28):9810-3. DOI: 10.1021/ja502904s. View

Liu P, Yang X, Sun S, Wang Q, Wang K, Huang J . Enzyme-free colorimetric detection of DNA by using gold nanoparticles and hybridization chain reaction amplification. Anal Chem. 2013; 85(16):7689-95. DOI: 10.1021/ac4001157. View

10.

Hou T, Li W, Liu X, Li F . Label-Free and Enzyme-Free Homogeneous Electrochemical Biosensing Strategy Based on Hybridization Chain Reaction: A Facile, Sensitive, and Highly Specific MicroRNA Assay. Anal Chem. 2015; 87(22):11368-74. DOI: 10.1021/acs.analchem.5b02790. View

11.

Zhang B, Liu B, Tang D, Niessner R, Chen G, Knopp D . DNA-based hybridization chain reaction for amplified bioelectronic signal and ultrasensitive detection of proteins. Anal Chem. 2012; 84(12):5392-9. DOI: 10.1021/ac3009065. View

12.

Yin P, Choi H, Calvert C, Pierce N . Programming biomolecular self-assembly pathways. Nature. 2008; 451(7176):318-22. DOI: 10.1038/nature06451. View

13.

Chen R, Blackstock D, Sun Q, Chen W . Dynamic protein assembly by programmable DNA strand displacement. Nat Chem. 2018; 10(4):474-481. DOI: 10.1038/s41557-018-0016-9. View

14.

Zhao R, Feng Z, Zhao Y, Jia L, Ma R, Zhang W . A sensitive electrochemical aptasensor for Mucin 1 detection based on catalytic hairpin assembly coupled with PtPdNPs peroxidase-like activity. Talanta. 2019; 200:503-510. DOI: 10.1016/j.talanta.2019.03.012. View

15.

Yan Y, Shen B, Wang H, Sun X, Cheng W, Zhao H . A novel and versatile nanomachine for ultrasensitive and specific detection of microRNAs based on molecular beacon initiated strand displacement amplification coupled with catalytic hairpin assembly with DNAzyme formation. Analyst. 2015; 140(16):5469-74. DOI: 10.1039/c5an00920k. View

16.

Liu R, Zhang S, Zheng T, Chen Y, Wu J, Wu Z . Intracellular Nonenzymatic Growth of Three-Dimensional DNA Nanostructures for Imaging Specific Biomolecules in Living Cells. ACS Nano. 2020; 14(8):9572-9584. DOI: 10.1021/acsnano.9b09995. View

17.

Zeng R, Zhang L, Luo Z, Tang D . Palindromic Fragment-Mediated Single-Chain Amplification: An Innovative Mode for Photoelectrochemical Bioassay. Anal Chem. 2019; 91(12):7835-7841. DOI: 10.1021/acs.analchem.9b01557. View

18.

Li C, Zhang J, Gao Y, Luo S, Wu Z . Nonenzymatic Autonomous Assembly of Cross-Linked Network Structures from Only Two Palindromic DNA Components for Intracellular Fluorescence Imaging of miRNAs. ACS Sens. 2022; 7(2):601-611. DOI: 10.1021/acssensors.1c02504. View