Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging.
Daniel EnglertEva-Maria BurgerFranziska GrünMrigank S VermaJens LacknerMarko LampeBastian BühlerJanin SchokolowskiGerd Ulrich NienhausAndres JäschkeMurat SunbulPublished in: Nature communications (2023)
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.
Keyphrases
- high resolution
- living cells
- single molecule
- gold nanoparticles
- sensitive detection
- fluorescent probe
- label free
- single cell
- small molecule
- deep learning
- fluorescence imaging
- oxidative stress
- cell therapy
- computed tomography
- quantum dots
- gene expression
- dna methylation
- molecular dynamics
- nucleic acid
- induced apoptosis
- convolutional neural network
- stem cells
- magnetic nanoparticles
- endoplasmic reticulum stress
- soft tissue
- breast reconstruction
- pet imaging
- genome wide association study
- pi k akt