Search of protein kinases responsible for phosphorylation of a major sigma factor in higher plants

Abstract

To avoid the generation of reactive oxygen species, makeup of PS II and PS I must be synchronized. We have found that a major sigma factor, SIG1, in RNA polymerase is phosphorylated in dim light to suppress the expression of photosynthesis genes, whereas SIG1 is dephosphorylated under irradiance to release the suppression.

We have employed a wheat germ cell-free protein production system in combination with AlphaScreen (PerkinElmer) to detect protein-protein interaction in vitro to search protein kinases responsible for phosphorylation of SIG1. SIG1 and approx. 800 protein kinases have been synthesized in vitro using RIKEN Arabidopsis Full-Length (RAFL) clones and subjected to the screening. We have obtained 49 candidate protein kinases. A phosphorylation assay has shown an activity to phosphorylate the Thr-170 of SIG1 in the wheat germ extracts.