Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging
Abstract Live-cell RNA imaging with high spatial and temporal resolution remains a major challenge. Here we report the development of RhoBAST:SpyRho, a fluorescent light-up aptamer (FLAP) system ideally suited for visualizing RNAs in live or fixed cells with various advanced fluorescence microscopy...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2023-06-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-023-39611-1 |
| _version_ | 1797789714881708032 |
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| author | Daniel Englert Eva-Maria Burger Franziska Grün Mrigank S. Verma Jens Lackner Marko Lampe Bastian Bühler Janin Schokolowski G. Ulrich Nienhaus Andres Jäschke Murat Sunbul |
| author_facet | Daniel Englert Eva-Maria Burger Franziska Grün Mrigank S. Verma Jens Lackner Marko Lampe Bastian Bühler Janin Schokolowski G. Ulrich Nienhaus Andres Jäschke Murat Sunbul |
| author_sort | Daniel Englert |
| collection | DOAJ |
| description | Abstract Live-cell RNA imaging with high spatial and temporal resolution remains a major challenge. Here we report the development of RhoBAST:SpyRho, a fluorescent light-up aptamer (FLAP) system ideally suited for visualizing RNAs in live or fixed cells with various advanced fluorescence microscopy modalities. Overcoming problems associated with low cell permeability, brightness, fluorogenicity, and signal-to-background ratio of previous fluorophores, we design a novel probe, SpyRho (Spirocyclic Rhodamine), which tightly binds to the RhoBAST aptamer. High brightness and fluorogenicity is achieved by shifting the equilibrium between spirolactam and quinoid. With its high affinity and fast ligand exchange, RhoBAST:SpyRho is a superb system for both super-resolution SMLM and STED imaging. Its excellent performance in SMLM and the first reported super-resolved STED images of specifically labeled RNA in live mammalian cells represent significant advances over other FLAPs. The versatility of RhoBAST:SpyRho is further demonstrated by imaging endogenous chromosomal loci and proteins. |
| first_indexed | 2024-03-13T01:54:36Z |
| format | Article |
| id | doaj.art-9497349644eb460a9fa92b4db02b0bfb |
| institution | Directory Open Access Journal |
| issn | 2041-1723 |
| language | English |
| last_indexed | 2024-03-13T01:54:36Z |
| publishDate | 2023-06-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj.art-9497349644eb460a9fa92b4db02b0bfb2023-07-02T11:19:32ZengNature PortfolioNature Communications2041-17232023-06-0114111310.1038/s41467-023-39611-1Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imagingDaniel Englert0Eva-Maria Burger1Franziska Grün2Mrigank S. Verma3Jens Lackner4Marko Lampe5Bastian Bühler6Janin Schokolowski7G. Ulrich Nienhaus8Andres Jäschke9Murat Sunbul10Institute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Applied Physics (APH), Karlsruhe Institute of Technology (KIT)Institute of Applied Physics (APH), Karlsruhe Institute of Technology (KIT)Advanced Light Microscopy Facility, European Molecular Biology LaboratoryInstitute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Applied Physics (APH), Karlsruhe Institute of Technology (KIT)Institute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityInstitute of Pharmacy and Molecular Biotechnology (IPMB), Heidelberg UniversityAbstract Live-cell RNA imaging with high spatial and temporal resolution remains a major challenge. Here we report the development of RhoBAST:SpyRho, a fluorescent light-up aptamer (FLAP) system ideally suited for visualizing RNAs in live or fixed cells with various advanced fluorescence microscopy modalities. Overcoming problems associated with low cell permeability, brightness, fluorogenicity, and signal-to-background ratio of previous fluorophores, we design a novel probe, SpyRho (Spirocyclic Rhodamine), which tightly binds to the RhoBAST aptamer. High brightness and fluorogenicity is achieved by shifting the equilibrium between spirolactam and quinoid. With its high affinity and fast ligand exchange, RhoBAST:SpyRho is a superb system for both super-resolution SMLM and STED imaging. Its excellent performance in SMLM and the first reported super-resolved STED images of specifically labeled RNA in live mammalian cells represent significant advances over other FLAPs. The versatility of RhoBAST:SpyRho is further demonstrated by imaging endogenous chromosomal loci and proteins.https://doi.org/10.1038/s41467-023-39611-1 |
| spellingShingle | Daniel Englert Eva-Maria Burger Franziska Grün Mrigank S. Verma Jens Lackner Marko Lampe Bastian Bühler Janin Schokolowski G. Ulrich Nienhaus Andres Jäschke Murat Sunbul Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging Nature Communications |
| title | Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging |
| title_full | Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging |
| title_fullStr | Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging |
| title_full_unstemmed | Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging |
| title_short | Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging |
| title_sort | fast exchanging spirocyclic rhodamine probes for aptamer based super resolution rna imaging |
| url | https://doi.org/10.1038/s41467-023-39611-1 |
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