Signatures of Antagonistic Pleiotropy in a Bacterial Flagellin Epitope.

Published: 4 March 2021| Version 1 | DOI: 10.17632/ccvvsv8g3w.1
Youssef Belkhadir


Immune sensors of higher organisms have been trained (over evolutionary time scales) to read molecular signals issued from, and required for, the maintenance of vital microbe functions. The genes encoding these signals are required for survival and, therefore, under strong negative selection. Otherwise, they would be eliminated through natural selection in order to evade host recognition. Despite detailed characterization of these systems over the last decades, relatively little is known about how co-evolutionary forces have influenced microbial fitness at the molecular level. There are very few systems where the tools and mechanistic details exist to address this properly. Here, we study the coevolutionary forces of non-self perception using Arabidopsis and Pseudomonads as reference systems. The model is outstanding because there are many genomic tools, mechanistic details are known for certain immune signaling pathways, and end-point assays can be performed on a full organism. In our case, we set out to understand how the plant immune sensor FLS2 (which shares structural similarities with TLR5) has influenced the evolution of flagellin (the main structural component of the bacterial flagellum) and, in turn, how this influenced the ability of Pseudomonas to move. In this study, we take full advantage of the rapid developments in proteomics-enabled chemistry and tested > 1000 physical interactions between FLS2 and mutated epitopes of flagellin. We also engineered > 400 synthetic Pseudomonas strains to assess the impact of epitope mutations on motility. We generated > 50 synthetic peptides to functionally validate our screen results in living plants. We discovered that antagonistic pleiotropy (the ability of a gene to show opposing effects in different context) have impacted molecularly the relationships between this flagellin epitope and FLS2 and, in turn, have balanced bacterial motility against FLS2 detection to promote stable colonization of Arabidopsis by bacterial communities. It is postulated that antagonistic pleiotropy acts to allow organisms phenotypic flexibility in different environments. Yet, little direct molecular evidence exists for its role in adaptation. Here we provide a clear-cut molecular example of this role. These results have broad implications to fully understand non-self perception in lower and higher metazoans, and also provide a guideline for next generation analyses aimed at understanding the molecular mechanisms of evolutionary trade-offs beyond the systems considered in this work. The methods we employed range from detailed biophysical characterization of receptor-epitope interactions, in vivo biochemistry, molecular genetics and algorithm analysis.



Gregor Mendel Institute of Molecular Plant Biology GmbH


Plant Biochemistry, Biophysical Methods, Bacterial Genetics, Epitope Mapping, Immune Receptor Binding