Affiliation:
1. Center for Desert Agriculture, Biological and Environmental Science & Engineering Division (BESE) King Abdullah University of Science and Technology (KAUST) Thuwal Saudi Arabia
2. Institute for Biophysical Chemistry, Hannover Medical School Carl‐Neuberg‐Straße 1 Hannover 30625 Germany
3. Max‐Planck‐Institute of Molecular Plant Physiology Am Muhlenberg 1 Potsdam‐Golm 14476 Germany
4. Core facility for Mass Spectrometry and Proteomics, Zentrum fur Molekulare Biologie der Universitat Heidelberg Im Neuenheimer Feld 329 Heidelberg 69120 Germany
5. School of BioSciences The University of Melbourne Parkville Victoria 3010 Australia
6. European Molecular Biology Laboratory (EMBL) Heidelberg Heidelberg 69117 Germany
7. Department of Plant Microbe Interactions Max Planck Institute for Plant Breeding Research 50829 Cologne Germany
8. Boyce Thompson Institute (BTI) Cornell University 533 Tower Rd. Ithaca New York 14853 USA
Abstract
SUMMARYIdentification of protein interactors is ideally suited for the functional characterization of small molecules. 3′,5′‐cAMP is an evolutionary ancient signaling metabolite largely uncharacterized in plants. To tap into the physiological roles of 3′,5′‐cAMP, we used a chemo‐proteomics approach, thermal proteome profiling (TPP), for the unbiased identification of 3′,5′‐cAMP protein targets. TPP measures shifts in the protein thermal stability upon ligand binding. Comprehensive proteomics analysis yielded a list of 51 proteins significantly altered in their thermal stability upon incubation with 3′,5′‐cAMP. The list contained metabolic enzymes, ribosomal subunits, translation initiation factors, and proteins associated with the regulation of plant growth such as CELL DIVISION CYCLE 48. To functionally validate obtained results, we focused on the role of 3′,5′‐cAMP in regulating the actin cytoskeleton suggested by the presence of actin among the 51 identified proteins. 3′,5′‐cAMP supplementation affected actin organization by inducing actin‐bundling. Consistent with these results, the increase in 3′,5′‐cAMP levels, obtained either by feeding or by chemical modulation of 3′,5′‐cAMP metabolism, was sufficient to partially rescue the short hypocotyl phenotype of the actin2 actin7 mutant, severely compromised in actin level. The observed rescue was specific to 3′,5′‐cAMP, as demonstrated using a positional isomer 2′,3′‐cAMP, and true for the nanomolar 3′,5′‐cAMP concentrations reported for plant cells. In vitro characterization of the 3′,5′‐cAMP–actin pairing argues against a direct interaction between actin and 3′,5′‐cAMP. Alternative mechanisms by which 3′,5′‐cAMP would affect actin dynamics, such as by interfering with calcium signaling, are discussed. In summary, our work provides a specific resource, 3′,5′‐cAMP interactome, as well as functional insight into 3′,5′‐cAMP‐mediated regulation in plants.
Funder
King Abdullah University of Science and Technology
Max-Planck-Gesellschaft
National Science Foundation
Subject
Cell Biology,Plant Science,Genetics
Cited by
4 articles.
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