Expanding the Toolkit of Fluorescent Biosensors for Studying Mitogen Activated Protein Kinases in Plants.

Mitogen-activated protein kinases (MAPKs) are key regulators of numerous biological processes in plants. To better understand the mechanisms by which these kinases function, high resolution measurement of MAPK activation kinetics in different biological contexts would be beneficial. One method to measure MAPK activation in plants is via fluorescence-based genetically-encoded biosensors, which can provide real-time readouts of the temporal and spatial dynamics of kinase activation in living tissue.

A Förster resonance energy transfer sensor for live-cell imaging of mitogen-activated protein kinase activity in Arabidopsis.

The catalytic activity of mitogen-activated protein kinases (MAPKs) is dynamically modified in plants. Since MAPKs have been shown to play important roles in a wide range of signaling pathways, the ability to monitor MAPK activity in living plant cells would be valuable. Here, we report the development of a genetically encoded MAPK activity sensor for use in Arabidopsis thaliana.

A simple microfluidic device for live cell imaging of Arabidopsis cotyledons, leaves, and seedlings.

One of the challenges of performing live-cell imaging in plants is establishing a system for securing the sample during imaging that allows for the rapid addition of treatments. Here we report how a commercially available device called a HybriWell can be repurposed to create an imaging chamber suitable for Arabidopsis seedlings, cotyledons and leaves. Liquid in the imaging chamber can be rapidly exchanged to introduce chemical treatments via microfluidic passive pumping.

Analysis of Phosphorylation of the Receptor-Like Protein Kinase HAESA during Arabidopsis Floral Abscission.

Receptor-like protein kinases (RLKs) are the largest family of plant transmembrane signaling proteins. Here we present functional analysis of HAESA, an RLK that regulates floral organ abscission in Arabidopsis. Through in vitro and in vivo analysis of HAE phosphorylation, we provide evidence that a conserved phosphorylation site on a region of the HAE protein kinase domain known as the activation segment positively regulates HAE activity.

Letting go is never easy: abscission and receptor-like protein kinases.

Abscission is the process by which plants discard organs in response to environmental cues/stressors, or as part of their normal development. Abscission has been studied throughout the history of the plant sciences and in numerous species. Although long studied at the anatomical and physiological levels, abscission has only been elucidated at the molecular and genetic levels within the last two decades, primarily with the use of the model plant Arabidopsis thaliana.

A simple in vitro method to measure autophosphorylation of protein kinases.

Receptor-like protein kinases (RLKs) are a large and important group of plant proteins involved in numerous aspects of development and stress response. Within this family, homo-oligermization of receptors followed by autophosphorylation of the intracellular protein kinase domain appears to be a widespread mechanism to regulate protein kinase activity. In vitro studies of several RLKs have identified autophosphorylation sites involved in regulation of catalytic activity and signaling in vivo.