Re-characterization of hammerhead ribozymes as molecular tools for intermolecular RNA cleavage

Liu, Getong; Huang, Xin; Pu, Qinlin; Zhao, You Xing; Du, Feng; Dong, Juan; Cui, Xin; Tang, Zhuo

doi:10.1039/c7ob00995j

Cited by 3 publications

(6 citation statements)

References 36 publications

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“…Furthermore, we investigated the adaptability of our sensor with other ribozymes by comparing the effects of different ribozymes on the sensor performance. The kinetics of the cleavage reaction on three kinds of trans-cleaving ribozymes through monitoring fluorescence signal under different reaction times. , The experimental results in Figure D,E revealed that the fluorescence signal generated by the enzyme digestion reaction gradually increased with the prolongation of reaction time. According to the kinetic curve of the catalytic reaction, the Michaelis constant and maximum reaction rate could be calculated, which is 3.67 ( K m )/1.02 ( V max ) for hammerhead ribozyme (II), 18.11 ( K m )/1.05 ( V max ) for hammerhead ribozyme (I), and 124.45 ( K m )/0.82 ( V max ) for HDV ribozyme, respectively.…”

Section: Resultsmentioning

confidence: 98%

“…The kinetics of the cleavage reaction on three kinds of trans-cleaving ribozymes through monitoring fluorescence signal under different reaction times. 25,26 The experimental results in Figure 3D,E of the catalytic reaction, the Michaelis constant and maximum reaction rate could be calculated, which is 3.67 (K m )/1.02 (V max ) for hammerhead ribozyme (II), 18.11 (K m )/1.05 (V max ) for hammerhead ribozyme (I), and 124.45 (K m )/0.82 (V max ) for HDV ribozyme, respectively. The results show that our sensor has good adaptability with other ribozymes and the catalytic efficiency of hammerhead ribozyme is better than that of HDV ribozyme.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

mentioning

confidence: 94%

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Cell-Free Biosensing Genetic Circuit Coupled with Ribozyme Cleavage Reaction for Rapid and Sensitive Detection of Small Molecules

Zhang

et al. 2023

ACS Synth. Biol.

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Synthetic biological systems have been utilized to develop a wide range of genetic circuits and components that enhance the performance of biosensing systems. Among them, cell-free systems are emerging as important platforms for synthetic biology applications. Genetic circuits play an essential role in cell-free systems, mainly consisting of sensing modules, regulation modules, and signal output modules. Currently, fluorescent proteins and aptamers are commonly used as signal outputs. However, these signal output modes cannot simultaneously achieve faster signal output, more accurate and reliable performance, and signal amplification. Ribozyme is a highly structured and catalytic RNA molecule that can specifically recognize and cut specific substrate sequences. Here, by adopting ribozyme as the signal output, we developed a cell-free biosensing genetic circuit coupled with the ribozyme cleavage reaction, enabling rapid and sensitive detection of small molecules. More importantly, we have also successfully constructed a 3D-printed sensor array and thereby achieved high-throughput analysis of an inhibitory drug. Furthermore, our method will help expand the application range of ribozyme in the field of synthetic biology and also optimize the signal output system of cell-free biosensing, thus promoting the development of cell-free synthetic biology in biomedical research, clinical diagnosis, environmental monitoring, and food inspection.

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Section: Resultsmentioning

confidence: 98%

Section: ■ Results and Discussionmentioning

confidence: 99%

mentioning

confidence: 94%

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Cell-Free Biosensing Genetic Circuit Coupled with Ribozyme Cleavage Reaction for Rapid and Sensitive Detection of Small Molecules

Zhang

et al. 2023

ACS Synth. Biol.

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“…Some researchers claimed this ribozyme could not cleave efficiently at GUG↓ site. They designed hammerhead ribozymes with a new cleaving site, "DWH" (D = A/U/ G, W = A/U, and H = A/U/C) and an optimal binding arm length of (8-9 nucleotides) to achieve trans-cleavage of a single-stranded RNA molecule [82].…”

Section: Deoxyribozymes and Ribozymesmentioning

confidence: 99%

“…Its expression was decreased 1000 Fig. 3 The secondary structures of hammerhead and engineered ribozymes [82] Fathi Dizaji Egyptian Journal of Medical Human Genetics (2020) 21:41…”

Section: Genome Engineering Toolsmentioning

confidence: 99%

Strategies to target long non-coding RNAs in cancer treatment: progress and challenges

Dizaji

2020

Egypt J Med Hum Genet

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Background: Long non-coding RNAs are important regulators of gene expression and diverse biological processes. Their aberrant expression contributes to a verity of diseases including cancer development and progression, providing them with great potential to be diagnostic and prognostic biomarkers and therapeutic targets. Therefore, they can have a key role in personalized cancer medicine. This review aims at introducing possible strategies to target long ncRNAs therapeutically in cancer. Also, chemical modification of nucleic acid-based therapeutics to improve their pharmacological properties is explained. Then, approaches for the systematic delivery of reagents into the tumor cells or organs are briefly discussed, followed by describing obstacles to the expansion of the therapeutics. Main text: Long ncRNAs function as oncogenes or tumor suppressors, whose activity can modulate all hallmarks of cancer. They are expressed in a very restricted spatial and temporal pattern and can be easily detected in the cells or biological fluids of patients. These properties make them excellent targets for the development of anticancer drugs. Targeting methods aim to attenuate oncogenic lncRNAs or interfere with lncRNA functions to prevent carcinogenesis. Numerous strategies including suppression of oncogenic long ncRNAs, alternation of their epigenetic effects, interfering with their function, restoration of downregulated or lost long ncRNAs, and recruitment of long ncRNAs regulatory elements and expression patterns are recommended for targeting long ncRNAs therapeutically in cancer. These approaches have shown inhibitory effects on malignancy. In this regard, proliferation, migration, and invasion of tumor cells have been inhibited and apoptosis has been induced in different cancer cells in vitro and in vivo. Downregulation of oncogenic long ncRNAs and upregulation of some growth factors (e.g., neurotrophic factor) have been achieved. Conclusions: Targeting long non-coding RNAs therapeutically in cancer and efficient and safe delivery of the reagents have been rarely addressed. Only one clinical trial involving lncRNAs has been reported. Among different technologies, RNAi is the most commonly used and effective tool to target lncRNAs. However, other technologies need to be examined and further research is essential to put lncRNAs into clinical practice.

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Chemically Cross-Linked Hammerhead Ribozyme as an Efficient RNA Interference Tool

Wang,

Wu,

Zhang

et al. 2024

J. Am. Chem. Soc.

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RNA-cleaving ribozymes are promising candidates as general tools of RNA interference (RNAi) in gene manipulation. However, compared with other RNA systems, such as siRNA and CRISPR technologies, the ribozyme tools are still far from broad applications on RNAi due to their poor performance in the cellular context. In this work, we report an efficient RNAi tool based on chemically modified hammerhead ribozyme (HHR). By the introduction of an intramolecular linkage into the minimal HHR to reconstruct the distal interaction within the tertiary ribozyme structure, this cross-linked HHR exhibits efficient RNA substrate cleavage activities with almost no sequence constraint. Cellular experiments suggest that both exogenous and endogenous RNA expression can be dramatically knocked down by this HHR tool with levels comparable to those of siRNA. Unlike the widely applied protein-recruiting RNA systems (siRNA and CRISPR), this ribozyme tool functions solely on RNA itself with great simplicity, which may provide a new approach for gene manipulation in both fundamental and translational studies.

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Re-characterization of hammerhead ribozymes as molecular tools for intermolecular RNA cleavage

Cited by 3 publications

References 36 publications

Cell-Free Biosensing Genetic Circuit Coupled with Ribozyme Cleavage Reaction for Rapid and Sensitive Detection of Small Molecules

Cell-Free Biosensing Genetic Circuit Coupled with Ribozyme Cleavage Reaction for Rapid and Sensitive Detection of Small Molecules

Strategies to target long non-coding RNAs in cancer treatment: progress and challenges

Chemically Cross-Linked Hammerhead Ribozyme as an Efficient RNA Interference Tool

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