Functional Imaging in living Plants - Cell Biology meets Physiology

Abstract

The last quarter of a century has brought major developments in the acquisition of images from plants through improvements in microscopy equipment, software and technique. Likewise, step changes in image analysis tools and the utilization and iterative redesign of fluorescent protein based markers and probes has revolutionized the ability of researchers to ask fundamental questions in cell biology and physiology. This research topic gives a snapshot of the current shape of the field in the plant sciences.The articles contributed to this research topic are indicative of the work emerging from the plant imaging community and cover, variously, the characterization of individual protein functions; localization and interactions; the use of physiological biosensors; spectroscopic techniques, which utilize autofluorescence of plant tissues and label-free approaches; developmental studies and software engineering. These reflect the broad areas in which imaging is currently being used to functionally dissect plant processes.A focus in this research topic is the quantitative imaging of fluorescent sensors to explore cell function.Förster resonance energy transfer (FRET) and how sensitized emission may be used for quantification in vivo imaging is reviewed by Müller et al. (2013) who discuss a set of methods that allows for the analysis of molecular interactions, in the light of recent developments in fluorescence microscopy, which have achieved higher spatial and temporal resolution as well as a much-improved sensitivity. A comprehensive overview of FRET imaging is given with a focus on fluorescent proteins and the procedure and analysis of sensitized emission, which allows for a fast and repetitive monitoring of FRET efficiencies as required for the investigation of dynamic plant cells.A perspective on the use of genetically encoded fluorescent biosensors (including FRET-based probes) in plants is given by Gjetting et al. (2013). The authors discuss the development of a rapidly growing repertoire of genetically encoded fluorescent sensors and how these developments have been a key driver for functional imaging over the last two decades as well as how new sensors have been adopted by plant biology and future opportunities. Autofluorescence from photosynthetic pigments and secondary metabolites, mounting techniques affecting physiological status, sensor silencing and plant specific compartments, such as the apoplast, are identified as technical burdens that can hamper straightforward sensor usage in plants. Plant-adjusted sensor design, such as the usage of new fluorescent protein variants, and imaging techniques, like fluorescent lifetime imaging (FLIM), are recognized as technical opportunities to advance in vivo sensing in plants. Biological promise comes from bespoke sensing approaches in which the sensor is matched current questions of plant metabolism, physiology and signaling, such as sugar homeostasis, hormone regulation and pH dynamics of acidic compartments.The development and properties of pH probes as one group...