Article

Temporal Analysis of Sucrose-induced Phosphorylation Changes in Plasma Membrane Proteins of Arabidopsis

Carnegie Institute, Stanford, California, United States
Molecular & Cellular Proteomics (Impact Factor: 6.56). 11/2007; 6(10):1711-26. DOI: 10.1074/mcp.M700164-MCP200
Source: PubMed

ABSTRACT

Sucrose is the main product of photosynthesis and the most common transport form of carbon in plants. In addition, sucrose is a compound that serves as a signal affecting metabolic flux and development. Here we provide first results of externally induced phosphorylation changes of plasma membrane proteins in Arabidopsis. In an unbiased approach, seedlings were grown in liquid medium with sucrose and then depleted of carbon before sucrose was resupplied. Plasma membranes were purified, and phosphopeptides were enriched and subsequently analyzed quantitatively by mass spectrometry. In total, 67 phosphopeptides were identified, most of which were quantified over five time points of sucrose resupply. Among the identified phosphorylation sites, the well described phosphorylation site at the C terminus of plasma membrane H(+)-ATPases showed a relative increase in phosphorylation level in response to sucrose. This corresponded to a significant increase of proton pumping activity of plasma membrane vesicles from sucrose-supplied seedlings. A new phosphorylation site was identified in the plasma membrane H(+)-ATPase AHA1 and/or AHA2. This phosphorylation site was shown to be crucial for ATPase activity and overrode regulation via the well known C-terminal phosphorylation site. Novel phosphorylation sites were identified for both receptor kinases and cytosolic kinases that showed rapid increases in relative intensities after short times of sucrose treatment. Seven response classes were identified including non-responsive, rapid increase (within 3 min), slow increase, and rapid decrease. Relative quantification of phosphorylation changes by phosphoproteomics provides a means for identification of fast responses to external stimuli in plants as a basis for further functional characterization.

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    • "Over the last decade, the application of welldeveloped proteomics and mass spectrometry techniques has led to a comprehensive description of co-translational and post-translational modifications of plant aquaporins (Santoni et al. 2006; Maurel 2007; Prak et al. 2008; Casado-Vela et al. 2010; Di Pietro et al. 2013). Phosphorylation of PIPs was specifically investigated in several species growing under various environmental conditions (Niittylä et al. 2007; Prak et al. 2008; Van Wilder et al. 2008; Kline et al. 2010; Di Pietro et al. 2013; Prado et al. 2013; Wu et al. 2013). In particular, PIPs of Arabidopsis showed multiple and interdependent phosphorylations at adjacent sites of their C-terminal tail (Prak et al. 2008), which were modulated by numerous stimuli including salt, oxidative stress or light (Di Pietro et al. 2013; Prado et al. 2013). "

    Full-text · Article · Jul 2015
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    • "Over the last decade, the application of welldeveloped proteomics and mass spectrometry techniques has led to a comprehensive description of co-translational and post-translational modifications of plant aquaporins (Santoni et al. 2006; Maurel 2007; Prak et al. 2008; Casado-Vela et al. 2010; Di Pietro et al. 2013). Phosphorylation of PIPs was specifically investigated in several species growing under various environmental conditions (Niittylä et al. 2007; Prak et al. 2008; Van Wilder et al. 2008; Kline et al. 2010; Di Pietro et al. 2013; Prado et al. 2013; Wu et al. 2013). In particular, PIPs of Arabidopsis showed multiple and interdependent phosphorylations at adjacent sites of their C-terminal tail (Prak et al. 2008), which were modulated by numerous stimuli including salt, oxidative stress or light (Di Pietro et al. 2013; Prado et al. 2013). "
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