Expression of the nascent RNA transcript is regulated by its interaction with a number of proteins. The misregulation of such interactions can often result in impaired cellular functions that can lead to cancer and a number of diseases. Thus, our understanding of RNA-protein interactions within the cellular context is essential for the development of novel diagnostic and therapeutic tools. While there are many in vitro methods that analyze RNA-protein interactions in vivo approaches are scarce. Here we established a method based on fluorescence resonance energy transfer (FRET), which we term RNA-binding mediated FRET (RB-FRET), which determines RNA-protein interaction inside cells and tested it on hnRNP H protein binding to its cognate RNA. Using two different approaches, we provide evidence that RB-FRET is sensitive enough to detect specific RNA-protein interactions in the cell, providing a powerful tool to study spatial and temporal localization of specific RNA-protein complexes.
- MeSH
- genetické vektory genetika MeSH
- HeLa buňky MeSH
- lidé MeSH
- proteiny vázající RNA genetika metabolismus MeSH
- rezonanční přenos fluorescenční energie metody MeSH
- RNA analýza metabolismus MeSH
- sekvence nukleotidů MeSH
- substrátová specifita MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- hodnotící studie MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH