RNA imaging in living cells - methods and applications

Abstract

Numerous types of transcripts perform multiple functions in cells, and these functions are mainly facilitated by the interactions of the RNA with various proteins and other RNAs. Insight into the dynamics of RNA biosynthesis, processing and cellular activities is highly desirable because this knowledge will deepen our understanding of cell physiology and help explain the mechanisms of RNA-mediated pathologies. In this review, we discuss the live RNA imaging systems that have been developed to date. We highlight information on the design of these systems, briefly discuss their advantages and limitations and provide examples of their numerous applications in various organisms and cell types. We present a detailed examination of one application of RNA imaging systems: this application aims to explain the role of mutant transcripts in human disease pathogenesis caused by triplet repeat expansions. Thus, this review introduces live RNA imaging systems and provides a glimpse into their various applications.

Keywords: RNA fluorescence imaging; RNA localization; RNA nuclear foci; live-cell imaging systems; triplet repeat diseases.

Figure 1.

Vector-based systems for RNA live imaging. Schematic structures of RNAs with protein partners or fluorescent dyes are presented (a–l). Additionally, examples of genetic constructs used for imaging experiments are depicted (o–r). (a) MS2 systems, (b) λN22 system, (c) BglG systems, (d) PP7 system, (e) U1Ap system, (f) HTLV-1 Rex system, (g) TAT system, (h) REV system, (i) eIF4A system, BiFC with the use of 2 domains, (j) Spinach system, (k) Malachite green system, (l) SRB-2 system, (m) BiFC with the use of 2 systems, (n) TriFC, (o) DNA construct for MS2 system, (p) DNA constructs for BiFC with 2 systems and (r) system for gene locus, mRNA and protein product imaging.

Figure 2.

RNA cellular journey in neurons. The figure presents RNA-related processes, which were investigated in neuronal cell lines using live imaging systems. Followed RNA and its aggregates are depicted in green, co-transported RNAs are in yellow and interacting proteins are in black. Protein products are illustrated in red, and gene loci are pink.