The cytosolic translation repressor NAB1 as a control switch for light-harvesting

Olaf Kruse, Jan H. Mussgnug & Lutz Wobbe

University of Bielefeld, Department of Biology, AlgaeBioTech Group, 33615 Bielefeld/Germany

The cytosolic RNA-binding protein NAB1 represses translation of LHCII encoding mRNAs by sequestration into translationally silent mRNP complexes in the green alga C. reinhardtii. Analysis of NAB1-RNA complex formation in vivo revealed that NAB1 recognizes LHCII transcripts with different affinities. Exposure of recombinant NAB1 to thiol oxidants and reductants reversibly altered the redox state of two cysteines in the C-terminal RRM domain of the protein. Oxidation of these cysteines in vitro resulted in intramolecular disulfide bridge formation and was accompanied by a strong decrease in RNA binding affinity. Re-reduction of oxidized NAB1 cysteines restored the binding affinity for the target mRNA sequence. In order to confirm the relevance of reversible NAB1 cysteine oxidation for the regulation of its activity in vivo we replaced both cysteines by serine applying site-directed mutagenesis. All examined cysteine single and double mutants exhibited a reduced antenna at PSII caused by a perturbed NAB1 deactivation mechanism. Comparative polysome analysis of LHCBM6 translation between a control strain and a mutant strain with a single cysteine mutation within NAB1 clearly demonstrated the correlation between the ability of disulfide formation and functional translational repression. Our new results demonstrated for the first time that cytosolic translation of nucleus encoded photosynthetic genes is redox-regulated via reversible cysteine modification of a transcript-specific translation repressor protein.

e-mail address of presenting author: olaf.kruse@uni-bielefeld.de