Methods for visualizing RNA processing and transport pathways in living cells

Recent advances in fluorescence microscopy, imaging, and probe technology p rovided possibilities to study the spatial and temporal distribution of RNA species in living cells. While some methods: have been developed to locali ze all nascent or poly (A) containing transcripts others have been develope d to study the in vivo distribution of specific RNA species. Irrespective o f the method that has been used, the results of these studies provided impo rtant information concerning the localization and the cellular transport pa thways of RNAs. Also, the picture emerges that RNA molecules travel through the nucleus at much faster speed, equaling that of free diffusion, than pr eviously anticipated. Still, a major challenge proves to be the development of a microscopic detection technique that allows specific, in vivo, detect ion of low levels of RNA species by fluorescence in situ hybridization, wit hout interfering fluorescent background signals derived from non-hybridized probe sequences and autofluorescent cell components. By applying photoacti vatable caged fluorochrome-, molecular beacon-, or fluorescence resonance e nergy transfer (FRET)-based detection methods an important step in the futu re of living cell analysis has already been made.