Detached barley leaves as a model for studying cytokinin control of plastid gene regulation

Abstract

We suggest a design of experiment, which permits studying a differential regulation of chloroplast gene transcription by exogenous cytokinin, using detached first barley leaves. Such experiments are of great interest for investigation of molecular mechanisms of the chloroplast biogenesis hormonal regulation. It was demonstrated that a cytokinin control of plastid genome transcription depended on the plant age, a source of detached leaves, chloroplast position in the leaf blade, and the intensity and duration of leaf illumination. Transcription regulation was studied for the first barley leaves detached from 9-day-old barley plants grown in soil. The leaves were preincubated for 24 h in the light (270 μmol/(m2 s) and then transferred for 3 h on water or cytokinin solution (benzyladenine, 2.2 × 10−5 M) under similar lighting condition. Chloroplasts were isolated from the most sensitive to cytokinin apical leaf part and used for run-on transcription. From 26 tested genes, cytokinin activated the rate of transcription of 19 genes (by 2 to 9 times as compared with control) and did not affect transcription of 7 genes. Cytokinin-sensitive genes encode proteins of photosynthesis and the components of the plastid genome expression system. The results obtained allow transition to more detail studying the mechanisms of hormonal regulation of chloroplast genome transcription.