Characterization of the promoter of rice plastid psaA-psaB-rps14 operon and the DNA-specific binding proteins.

Transcriptional analysis of the promoter of rice psaA-psaB-rps14 operon was performed with Bal 31 deleted mutants in spinach chloroplast extracts. Two functional promoters in this operon (denoted as "-175" and "-129" promoters) were revealed to be utilized by the spinach transcription machinery; however only the "-175" promoter is utilized in vivo. The existence of psaA promoter-specific binding factors in the chloroplast extracts was indicated by competitive gel retardation assay and exonuclease III protection analysis. After ammonium sulfate fractionation of the extracts followed by heparin-agarose column chromatography, the sequence-specific binding factors were characterized to be 66-kDa and 31-kDa polypeptides. Southwestern analysis demonstrated that the two protein factors interact independently with the psaA promoter-containing DNA fragment.