Quantitative and functional posttranslational modification proteomics reveals that TREPH1 plays a role in plant touch-delayed bolting

Abstract

Plants can sense both intracellular and extracellular mechanical forces and can respond through morphological changes. The signaling components responsible for mechanotransduction of the touch response are largely unknown. Here, we performed a high-throughput SILIA (stable isotope labeling in Arabidopsis)-based quantitative phosphoproteomics analysis to profile changes in protein phosphorylation resulting from 40 seconds of force stimulation in Arabidopsis thaliana. Of the 24 touch-responsive phosphopeptides identified, many were derived from kinases, phosphatases, cytoskeleton proteins, membrane proteins and ion transporters. TOUCH-REGULATED PHOSPHOPROTEIN1 (TREPH1) and MAP KINASE KINASE 2 (MKK2) and/or MKK1 became rapidly phosphorylated in touch-stimulated plants. Both TREPH1 and MKK2 are required for touch-induced delayed flowering, a major component of thigmomorphogenesis. The treph1-1 and mkk2 mutants also exhibited defects in touch-inducible gene expression. A nonphosphorylatable site-specific isoform of TREPH1 (S625A) failed to restore touch-induced flowering delay of treph1-1, indicating the necessity of S625 for TREPH1 function and providing evidence consistent with the possible functional relevance of the touch-regulated TREPH1 phosphorylation. Bioinformatic analysis and biochemical subcellular fractionation of TREPH1 protein indicate that it is a soluble protein. Altogether, these findings identify new protein players in Arabidopsis thigmomorphogenesis regulation, suggesting that protein phosphorylation may play a critical role in plant force responses.Like neural systems in animals, plants respond to a delicate force signal, such as a light touch, with extreme sensitivity, being it thigmotropism, thigmonastic movement, and thigmomorphogenesis. To understand the complex force signaling networks in plants, we applied SILIA-based quantitative PTM proteomics to measure 40 seconds of protein phosphorylation changes in Arabidopsis in response to cotton and human hair touches, and identified 4895 repeatable and non-redundant phosphopeptides, 579 of which are novel phosphosites derived from the 509 phosphoprotein groups, and finally identified 24 TOUCH-REGUALTED PHOSPHOPROTEIN (TREPHs) groups. Consequent molecular biological and bioinformatic studies revealed that both TREPH1 and MKK2 proteins are required for Arabidopsis touch response. These studies suggest that protein phosphorylation is involved in mechanotransduction of plant thigmomorphogenesis.

A MYC2/MYC3/MYC4-dependent transcription factor network regulates water spray-responsive gene expression and jasmonate levels

“…Accordingly, proteomic analysis revealed that the abundance of over 300 proteins was altered in at least 1 time point after water spray. Several kinases/phosphatases were identified, confirming the importance of phosphorylation cascades in mechanical stimulation signaling (50). In addition, redox status seems to play an important role with many peroxidases, thioredoxins, and glutaredoxins being altered in abundance after water spray.…”

“…Transcriptional, posttranscriptional, and posttranslational mechanisms underlying the touch response have been identified. These include the identification of cis-regulatory regions, the characterization of active mRNA degradation components, and posttranslational modifications that affect touch-induced transcript accumulation levels (46)(47)(48)(49)(50). Although transcription factors are central to such transcriptional reprogramming, a regulatory network underlying the touch response remains to be identified.…”

AtPiezo Plays an Important Role in Root Cap Mechanotransduction

“…These genes are mainly encoding proteins related to calcium signals, enzymes related to pectin and cellulose biosynthesis, and kinases involved in plant disease resistance [ 6 , 7 ]. A recent report showed that TRHEP1, a cytoplasmic protein that can be phosphorylated, is involved in the process of flowering delay in response to touch treatment [ 8 ].…”

Functional analysis tools for post‐translational modification: a post‐translational modification database for analysis of proteins and metabolic pathways

“…Additionally, we obtained data supporting ubiquitinylation, SUMOylation, N ‐glycosylation, O ‐GlcNAcylation, acylation, S ‐nitrosylation and lysine acetylation events at 16 094 sites in 7238 proteins identified in large‐scale proteomic studies from the literature (Miller et al ., , ; Hemsley et al ., ; Kim et al ., ; Svozil et al ., ; Hu et al ., ; Johnson and Vert, ; Walton et al ., ; Xu et al ., , ; Koskela et al ., ; Liu et al ., ; Majeran et al ., ; Rytz et al ., ; Zeng et al ., ) (Table ). We also collected data supporting 19 043 mapped phosphorylation sites in 5783 proteins resulting from quantitative phosphoproteomics studies (Benschop et al ., ; Nühse et al ., ; Mithoe et al ., ; Zhang et al ., , ; Chen and Hoehenwarter, ; Minkoff et al ., ; Roitinger et al ., ; Cho et al ., ; Bhaskara et al ., ; Wang et al ., ) (Table ). To our knowledge, this is the most complete repository of Arabidopsis PTMs to date.…”