DOI: 10.5061/dryad.b8gtht7k4
title: Data from: A cucumber protein, Phloem Phosphate Stress Repressed 1, rapidly degrades in response to a phosphate stress condition

description: Under depleted external phosphate (Pi), many plant species adapt to this
 stress by initiating downstream signalling cascades. In plants, the
 vascular system delivers nutrients and signalling agents to control
 physiological and developmental processes. Currently, limited information
 is available regarding the direct role of phloem-borne long-distance
 signals in plant growth and development under Pi-stress conditions. Here,
 we report on the identification and characterization of a cucumber
 protein, Cucumis sativus Phloem Phosphate-Stress-Repressed 1 (CsPPSR1),
 whose level in the phloem translocation stream rapidly responds to imposed
 Pi-limiting conditions. CsPPSR1 degradation is mediated by the 26S
 proteasome; under Pi-sufficient conditions, CsPPSR1 is stabilized by its
 phosphorylation, within the sieve tube system, through the action of
 CsPPSR1 Kinase. Further, we discovered that CsPPSR1 Kinase was susceptible
 to Pi-starvation-induced degradation, in the sieve tube system. Our
 findings offer insight into a molecular mechanism underlying the response
 of phloem-borne proteins to Pi-limited stress conditions.
description: In-gel kinase assays were performed as follows: Briefly,
 <em>E. coli</em>-purified GFP and CsPPSR1-4M8H were used as
 substrates. Fast protein liquid chromatography (FPLC) fractionated
 cucumber phloem sap proteins were separated on a substrate-containing
 SDS-PAGE (200 µg/ml of each substrate). Conditioned gels were incubated
 with 50 µM ATP and [γ32P]-ATP (20 Ci/mL) for in-gel phosphorylation. The
 reaction was terminated by incubating the gels with gel wash solution (5%
 TCA [V/V] and 1% [W/V] sodium pyrophosphate) and phosphorimaging using a
 Storm 860 system (GE Healthcare Lifesciences). To identify the kinase that
 phosphorylated CsPPSR1, named CsPPSRK, fractions with CsPPSR1-specific
 phosphorylation activity were separated on a 13% SDS-PAGE gel. Then the
 approximate 40 kDa protein band with phosphorylation activity was excised
 for mass spectrometry analysis. These mass spectral data were interrogated
 to identify sequences associated with a protein kinase of the appropriate
 mass.
description: # A cucumber protein, Phloem Phosphate Stress Repressed 1, rapidly
 degrades in response to a phosphate stress condition
 Specific kinase to
 phosphorylate Phloem Phosphate Stress Repressed 1 (CsPPSR1) was identified
 through in-gel kinase assay. The SDS-PAGE resolved FPLC fraction that
 contains the specific signal to phosphorylate CsPPSR1 as substrate was
 analyzed by LC-MS/MS and the raw data sets were deposited into DRYAD. ##
 Description of the data and file structure The MASCOT file is
 PPAN1314.mzid.gz_GPM00500046882.MGF. The Spectra raw information is in
 PPAN1314.mzid. PPAN1314.mzid.gz_GPM00500046882.MGF file was used to encode
 mass spectra results; multiple spectra were encoded in one MGF. Every
 spectrum begins with the BEGINS IONS assessment and finishes with END
 IONS. PEPMASS is the precursor mass of the representative spectra. This
 MGF file describes Ions information within two columns: the first column
 is the ion mass (Unit: Dalton, Da), the second column is the ion charge
 (Unit: m/z, referred to as the mass-to-charge ratio of an ion).
 PPAN1314.mzid.gz_GPM00500046882.MGF and PPAN1314.mzid are raw data for
 in-gel kinase assay by using CsPPSR1 as a specific substrate. Figure S4
 presented the original gel image where the specific kinase signal was
 identified. ## Code/Software Scaffold software (licensed version) can be
 used to load the raw datasets.