Traditional molecular biology tools have elucidated the identities and functions of RNA molecules,which are essential to the understanding of gene transcription and protein translation.Deepening this research field wo...Traditional molecular biology tools have elucidated the identities and functions of RNA molecules,which are essential to the understanding of gene transcription and protein translation.Deepening this research field would further require the direct visualization of RNA dynamics such as the DNA-RNA interactions and RNA-protein interactions.Towards this goal,the rise of RNA imaging tools over the past 15 years has reformed how we looked at these processes.In this Emerging Topic,we first highlighted recent advances on three main RNA imaging tools based on the species of interacting molecules:RNA-RNA pairing,RNA-protein binding,and small molecule-RNA complex.We introduced the advantages of these tools from a technical viewpoint,including binding affinity,fluorescent turn-on ratio,stability,and impacts on targeted RNA.Next,we discussed new rising opportunities and future directions,echoing the state-of-the-art imaging tools in the fields of fluorescent proteins and small fluorescent molecules.Together,we believe this emerging field will bring new insights on how we study RNA biology in living systems.展开更多
RNA imaging in live cells can provide comprehensive information on the expression,localization,degradation,storage,and regulation of RNA in cells,which is crucial for basic biology and clinical research.Our previous r...RNA imaging in live cells can provide comprehensive information on the expression,localization,degradation,storage,and regulation of RNA in cells,which is crucial for basic biology and clinical research.Our previous research finds that slicing the facilitated crRNA in the typical CRISPR/Cas12a system at a fitted site did not affect its trans-cleavage activity,which was previously reported to be triggered by targeted ssDNA or dsDNA,and a mini crRNA-mediated CRISPR/Cas12a(MCM-CRISPR/Cas12a)system was proposed.Here,we further apply it to enhanced imaging of MicroRNAs in cells by designing the activator in the system as a molecular beacon(MB),which can form a hybrid double-stranded structure of DNA/RNA with the targeted MicroRNA.When targeted MicroRNA is present,the hairpin structure of the MB is opened and the system emits fluorescence.Simultaneously,the DNA-RNA formed by the targeted MicroRNA and MB activates the trans-cleavage activity of LbCas12a,partially cleaving the single-stranded DNA extended from the MB and further enhancing the fluorescence intensity of the system.We designed the MCM-CRISPR/Cas12a system for miRNA-21,miRNA-155,and miRNA-10b and successfully applied it for sensitive and specific imaging of these MicroRNAs both inside and outside cells.This study provides a new idea for the sensitive and specific imaging of multiple MicroRNAs within cells,which is important for studying the distribution and dynamic changes of MicroRNAs in cells.展开更多
RNA molecules are widely regarded as one of the most ancient molecular forms of life.In 1960s,Francis Crick,Leslie Orgel and Carl R.Woese proposed that RNAs might be the original biomolecule,a concept that later cryst...RNA molecules are widely regarded as one of the most ancient molecular forms of life.In 1960s,Francis Crick,Leslie Orgel and Carl R.Woese proposed that RNAs might be the original biomolecule,a concept that later crystallized into the“Primordial RNA World”theory.The discovery of catalytic RNAs,called ribozymes,in the 1980s provided compelling experimental support for this idea.Over billions of years of evolution,from simple to complex organisms,RNAs have maintained their central roles in both the expression and the regulation of genetic information.展开更多
As the most abundant biological entities with incredible diversity,bacteriophages(also known as phages)have been recognized as an important source of molecular machines for the development of genetic-engineering tools...As the most abundant biological entities with incredible diversity,bacteriophages(also known as phages)have been recognized as an important source of molecular machines for the development of genetic-engineering tools.At the same time,phages are crucial for establishing and improving basic theories of molecular biology.Studies on phages provide rich sources of essential elements for synthetic circuit design as well as powerful support for the improvement of directed evolution platforms.Therefore,phages play a vital role in the development of new technologies and central scientific concepts.After the RNA world hypothesis was proposed and developed,novel biological functions of RNA continue to be discovered.RNA and its related elements are widely used in many fields such as metabolic engineering and medical diagnosis,and their versatility led to a major role of RNA in synthetic biology.Further development of RNA-based technologies will advance synthetic biological tools as well as provide verification of the RNA world hypothesis.Most synthetic biology efforts are based on reconstructing existing biological systems,understanding fundamental biological processes,and developing new technologies.RNA-based technologies derived from phages will offer abundant sources for synthetic biological components.Moreover,phages as well as RNA have high impact on biological evolution,which is pivotal for understanding the origin of life,building artificial life-forms,and precisely reprogramming biological systems.This review discusses phage-derived RNA-based technologies terms of phage components,the phage lifecycle,and interactions between phages and bacteria.The significance of RNA-based technology derived from phages for synthetic biology and for understanding the earliest stages of biological evolution will be highlighted.展开更多
基金supports from National Key R&D Pro-gram of China(Grant 2022YFA1305400)National Natural Sci-ence Foundation of China(Grant 22104113,Grant 22274122).
文摘Traditional molecular biology tools have elucidated the identities and functions of RNA molecules,which are essential to the understanding of gene transcription and protein translation.Deepening this research field would further require the direct visualization of RNA dynamics such as the DNA-RNA interactions and RNA-protein interactions.Towards this goal,the rise of RNA imaging tools over the past 15 years has reformed how we looked at these processes.In this Emerging Topic,we first highlighted recent advances on three main RNA imaging tools based on the species of interacting molecules:RNA-RNA pairing,RNA-protein binding,and small molecule-RNA complex.We introduced the advantages of these tools from a technical viewpoint,including binding affinity,fluorescent turn-on ratio,stability,and impacts on targeted RNA.Next,we discussed new rising opportunities and future directions,echoing the state-of-the-art imaging tools in the fields of fluorescent proteins and small fluorescent molecules.Together,we believe this emerging field will bring new insights on how we study RNA biology in living systems.
基金supported by the Young Scientists Fund of the National Natural Science Foundation of China(No.32301155)Natural Science Foundation of Chongqing,China(CSTB2023NSCQ-MSX0388)+1 种基金Science and Technology Innovation Enhancement Project of Army Medical University(2022XQN33)Young Doctoral Talent Incubation Program of the Xinqiao Hospital,Army Military Medical University(2022YQB065).
文摘RNA imaging in live cells can provide comprehensive information on the expression,localization,degradation,storage,and regulation of RNA in cells,which is crucial for basic biology and clinical research.Our previous research finds that slicing the facilitated crRNA in the typical CRISPR/Cas12a system at a fitted site did not affect its trans-cleavage activity,which was previously reported to be triggered by targeted ssDNA or dsDNA,and a mini crRNA-mediated CRISPR/Cas12a(MCM-CRISPR/Cas12a)system was proposed.Here,we further apply it to enhanced imaging of MicroRNAs in cells by designing the activator in the system as a molecular beacon(MB),which can form a hybrid double-stranded structure of DNA/RNA with the targeted MicroRNA.When targeted MicroRNA is present,the hairpin structure of the MB is opened and the system emits fluorescence.Simultaneously,the DNA-RNA formed by the targeted MicroRNA and MB activates the trans-cleavage activity of LbCas12a,partially cleaving the single-stranded DNA extended from the MB and further enhancing the fluorescence intensity of the system.We designed the MCM-CRISPR/Cas12a system for miRNA-21,miRNA-155,and miRNA-10b and successfully applied it for sensitive and specific imaging of these MicroRNAs both inside and outside cells.This study provides a new idea for the sensitive and specific imaging of multiple MicroRNAs within cells,which is important for studying the distribution and dynamic changes of MicroRNAs in cells.
文摘RNA molecules are widely regarded as one of the most ancient molecular forms of life.In 1960s,Francis Crick,Leslie Orgel and Carl R.Woese proposed that RNAs might be the original biomolecule,a concept that later crystallized into the“Primordial RNA World”theory.The discovery of catalytic RNAs,called ribozymes,in the 1980s provided compelling experimental support for this idea.Over billions of years of evolution,from simple to complex organisms,RNAs have maintained their central roles in both the expression and the regulation of genetic information.
基金This work was financially supported by National Key Research and Development Project of China(Grant No.2018YFA0900103)International Cooperation and Exchange of the National Natural Science Foundation of China(Grant No.31961133019)National Natural Science Foundation of China(Grant No.31670991).
文摘As the most abundant biological entities with incredible diversity,bacteriophages(also known as phages)have been recognized as an important source of molecular machines for the development of genetic-engineering tools.At the same time,phages are crucial for establishing and improving basic theories of molecular biology.Studies on phages provide rich sources of essential elements for synthetic circuit design as well as powerful support for the improvement of directed evolution platforms.Therefore,phages play a vital role in the development of new technologies and central scientific concepts.After the RNA world hypothesis was proposed and developed,novel biological functions of RNA continue to be discovered.RNA and its related elements are widely used in many fields such as metabolic engineering and medical diagnosis,and their versatility led to a major role of RNA in synthetic biology.Further development of RNA-based technologies will advance synthetic biological tools as well as provide verification of the RNA world hypothesis.Most synthetic biology efforts are based on reconstructing existing biological systems,understanding fundamental biological processes,and developing new technologies.RNA-based technologies derived from phages will offer abundant sources for synthetic biological components.Moreover,phages as well as RNA have high impact on biological evolution,which is pivotal for understanding the origin of life,building artificial life-forms,and precisely reprogramming biological systems.This review discusses phage-derived RNA-based technologies terms of phage components,the phage lifecycle,and interactions between phages and bacteria.The significance of RNA-based technology derived from phages for synthetic biology and for understanding the earliest stages of biological evolution will be highlighted.
