Genetically encodable fluorescent biosensors for tracking signaling dynamics in living cells.

Fluorescence has long been recognized as a powerful tool for probing biological structure and function. Nearly sixty-five years ago, Coons and colleagues introduced the first fluorescent-antibody techniques as a means of detecting pneumococcal antigens in mouse tissue.1 In the years that followed, studies have been conducted in hundreds of cellular contexts using immunofluorescence and related techniques such as fluorescence in situ hybridization.2 Together, these studies have provided cell biologists with a unique glimpse into the molecular organization of cells and tissues. However, because such approaches typically require fixation prior to detection, they provide only a snapshot of the highly dynamic cellular processes that govern cell physiology at the molecular level. To add a crucial temporal component to the study of cell biology, researchers have developed a diverse set of genetically-encodable biosensors designed to probe dynamic cellular events in living cells with high spatial and temporal resolution. These sensors, which generally involve the incorporation of a fluorescent tag into the primary amino acid sequence of a protein or a selected protein domain, have enabled researchers to track various components of intracellular signaling networks in real time within the native cellular environment. In this review, we will first explore the molecular basis of several genetically-targetable fluorescent tags amenable to live cell imaging. We will then turn our attention to the development and use of fluorescent biosensors for studying dynamic signaling processes in living cells.