A novel plant protein undergoing light-induced phosphorylation and release from the photosynthetic thylakoid membranes

^†Division of Cell Biology, Linköping University, SE-58185 Linköping, Sweden; ^*Department of Biochemistry and Biophysics, Arrhenius Laboratories of Natural Sciences, Stockholm University, SE-10691 Stockholm, Sweden; ^‡Department of Medical Nutrition and Biosciences, Karolinska Institute, Novum, SE-14186 Huddinge, Sweden; and ^§Department of Biochemistry, Umeå University, SE-90187 Umeå, Sweden

Edited by George H. Lorimer, University of Maryland, College Park, MD, and approved November 22, 2002 (received for review September 8, 2002)

Abstract

The characteristics of a phosphoprotein with a relative electrophoretic mobility of 12 kDa have been unknown during two decades of studies on redox-dependent protein phosphorylation in plant photosynthetic membranes. Digestion of this protein from spinach thylakoid membranes with trypsin and subsequent tandem nanospray-quadrupole-time-of-flight mass spectrometry of the peptides revealed a protein sequence that did not correspond to any previously known protein. Sequencing of the corresponding cDNA uncovered a gene for a precursor protein with a transit peptide followed by a strongly basic mature protein with a molecular mass of 8,640 Da. Genes encoding homologous proteins were found on chromosome 3 of Arabidopsis and rice as well as in ESTs from 20 different plant species, but not from any other organisms. The protein can be released from the membrane with high salt and is also partially released in response to light-induced phosphorylation of thylakoids, in contrast to all other known thylakoid phosphoproteins, which are integral to the membrane. On the basis of its properties, this plant-specific protein is named thylakoid soluble phosphoprotein of 9 kDa (TSP9). Mass spectrometric analyses revealed the existence of non-, mono-, di-, and triphosphorylated forms of TSP9 and phosphorylation of three distinct threonine residues in the central part of the protein. The phosphorylation and release of TSP9 from the photosynthetic membrane on illumination favor participation of this basic protein in cell signaling and regulation of plant gene expression in response to changing light conditions.

Footnotes

↵ ¶ To whom correspondence should be addressed. E-mail: aleve{at}ibk.liu.se.
This paper was submitted directly (Track II) to the PNAS office.
Data deposition: The sequence reported in this paper has been deposited in the GenBank database (accession no. AJ507430).
Abbreviations:

PSII,

photosystem II;

CP43,

chlorophyll a binding protein of PSII;

D1 and D2,

photosystem II reaction center proteins;

FTR,

ferredoxin-thioredoxin reductase;

MALDI,

matrix-assisted laser desorption/ionization;

TOF,

time-of-flight;

TSP9,

thylakoid soluble phosphoprotein of 9 kDa