01/10/2018

An extracellular network map of Arabidopsis LRR receptor kinases demonstrates that they operate as unified regulatory network

In a landmark study, an international consortium of laboratories (3 groups from Austria, 2 from Canada, 2 from the USA, and 1 from the UK) spearheaded by Youssef Belkhadir at the Gregor Mendel Institute of Molecular Plant Biology of the Austrian Academy of Sciences, have released a comprehensive interaction network map of one of the most important classes of receptors, the leucine-rich repeat receptor kinases or LRR-RKs. The work entitled “An extracellular network of Arabidopsis leucine-rich repeat receptor kinases,” was published today as an advance online publication in the journal NATURE.

In both plants and animals, a family of proteins named receptor kinases are largely responsible for sensing the environment.  Plants use receptor kinases as ‘molecular antennas’ to recognize chemical signals, such as growth hormones or portions of proteins from pathogens, from outside the cell and initiate responses to this signal inside the cell.   The model plant Arabidopsis thaliana contains over 600 of these receptor kinases – ten times more than are found in humans -  and they are critical for plant growth, development, immunity, and stress response. Despite this importance, the function of only a handful of these proteins is known; even less is known about how these receptors interact with each to coordinate responses to often conflicting signals.

The lead laboratory directed by Dr. Belkhadir implemented a high-throughput assay to test interactions between the receptors in a pairwise manner. They cloned and expressed more than 400 extracellular domains of LRR-RKs and performed 40,000 interaction tests, testing whether each protein interacted with all of the others, and produced an interaction map displaying how the receptor kinases interact with one another. With their strong expertise in experimental systems biology and computational network science, the laboratories of the co-senior authors Prof. David Guttman and Prof. Darrell Desveaux, at the University of Toronto (Canada) analyzed the receptor interaction map using various algorithms, generating diverse hypotheses that were subsequently validated in the Belkhadir and Zipfel laboratories. The group of Prof. Cyril Zipfel at The Sainsbury Laboratory (TSL)  in Norwich (UK) provided its unique and world-renowned know-how for tackling challenging molecular mechanisms. Finally, the laboratory of Prof. Shahid Mukhtar at the University of Alabama at Birmingham (USA) provided a finely-tuned interaction mapping pipeline for interrogating interactions between the intracellular domains of these receptors.

“This work is the result of an outstanding team-effort that required a unique breadth of expertise” says Dr. Belkhadir. “Beyond this publication, the work showcases how collaborative approaches in life science can break barriers and push technical boundaries to address important biological questions… this is the most remarkable aspect for me” says Dr. Belkhadir.

 “After we generated this map, there were two major surprises” says Dr. Adam Mott a co-lead author. “First, LRR-RKs that have small extracellular domains interact with other LRR-RKs more often than their counterparts with large domains.  This suggests that these small LRR-RKs have evolved to coordinate the actions of the rest of the receptors.  Second, we identified several unknown LRR-RKs that appear to be critical for network integrity.”

They named the most important of these new LRR-RKs “APEX”, and its removal was predicted to cause the most severe disruptions to the rest of the network. Indeed, when APEX was removed along with other known LRR-RKs, the resulting plants were developmentally impaired.  According to Dr. Elwira Smakowska, the first lead author of the study, “We found that the absence of APEX led to significant changes in plant developmental and immune responses controlled by two other LRR-RKs that do not interact with APEX, and are actually several network steps away.  This demonstrates that our interaction and receptor network map can be mined to find important receptors that can affect multiple, diverse plant response pathways.”

According to Dr. Belkhadir, “This is the first big step in elucidating the network properties of receptor kinases and this step has changed the way we think about how these molecular antennas function; Interaction  datasets for these types of proteins are extremely challenging to produce and only a very few institutions in the world have the required know-how to do this… my lab is currently expanding this work to the other receptor classes in Arabidopsis, and moving out of this model system and into crops. Our ultimate goal is to identify important receptor kinases that can modify plant stress responses to make them more resistant to environmental stress, such as global warming, and pathogens.