Single Molecule Fluorescence Approaches Shed Light on Intracellular RNAs

Numerous RNAs that are found in particularly the eukaryotic cell and a detailed summary of the single molecule tools available to study RNAs intracellularly is reviewed. The majority of RNAs found in the eukaryotic cell do not code for protein but rather regulate gene expression via post-transcriptional gene silencing or epigenetic gene regulation. Intracellular single molecule techniques offer an unparalleled means to investigate the behavior of these emerging classes of non-coding RNAs (ncRNAs). Many RNA elements have been found to originate from overlapping loci, suggesting that similar RNA sequences can be distinctly generated or processed to perform different biological functions. RNA-seq technologies coupled with RNA-protein cross-linking have been successful in mapping RNA target binding sites of RNA-binding proteins on a genomic scale. miRNAs are the most ubiquitous small ncRNA in humans, with over 1500 different mammalian miRNA sequences discovered that represent more than 1% of the entire genome and thus the largest gene family.