Systematic <i>In Vivo</i> Characterization of Fluorescent Protein Maturation in Budding Yeast

Abstract

Fluorescent protein (FP) maturation can limit the accuracy with which dynamic intracellular processes are captured and reduce the <i>in vivo</i> brightness of a given FP in fast-dividing cells. The knowledge of maturation timescales can therefore help users determine the appropriate FP for each application. However, <i>in vivo</i> maturation rates can greatly deviate from <i>in vitro</i> estimates that are mostly available. In this work, we present the first systematic study of <i>in vivo</i> maturation for 12 FPs in budding yeast. To overcome the technical limitations of translation inhibitors commonly used to study FP maturation, we implemented a new approach based on the optogenetic stimulations of FP expression in cells grown under constant nutrient conditions. Combining the rapid and orthogonal induction of FP transcription with a mathematical model of expression and maturation allowed us to accurately estimate maturation rates from microscopy data in a minimally invasive manner. Besides providing a useful resource for the budding yeast community, we present a new joint experimental and computational approach for characterizing FP maturation, which is applicable to a wide range of organisms.