A proteomics approach identifies novel proteins involved in gravitropic signal transduction

Abstract

Premise: Plant organs use gravity as a guide to direct their growth. And although gravitropism has been studied since the time of Darwin, the mechanisms of signal transduction, those that connect the biophysical stimulus perception and the biochemical events of the response, are still not understood.

Methods: A quantitative proteomics approach was used to identify key proteins during the early events of gravitropism. Plants were subjected to a gravity persistent signal (GPS) treatment, and proteins were extracted from the inflorescence stem at early time points after stimulation. Proteins were labeled with isobaric tags for relative and absolute quantification (iTRAQ) reagents. Proteins were identified and quantified as a single step using tandem mass-spectrometry (MS/MS). For two of the proteins identified, mutants with T-DNA inserts in the corresponding genes were evaluated for gravitropic phenotypes.

Key results: A total of 82 proteins showed significant differential quantification between treatment and controls. Proteins were categorized into functional groups based on gene ontology terms and filtered using groups thought to be involved in the signaling events of gravitropism. For two of the proteins selected, GSTF9 and HSP81-2, knockout mutations resulted in defects in root skewing, waving, and curvature as well as in the GPS response of inflorescence stems.

Conclusion: Combining a proteomics approach with the GPS response, 82 novel proteins were identified to be involved in the early events of gravitropic signal transduction. As early as 2 and 4 min after a gravistimulation, significant changes occur in protein abundance. The approach was validated through the analysis of mutants exhibiting altered gravitropic responses.