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Nucleic Acids Research, 2003, Vol. 31, No. 24 7090-7098
© 2003 Oxford University Press

Article

Molecular genetic analysis of chloroplast gene promoters dependent on SIG2, a nucleus-encoded sigma factor for the plastid-encoded RNA polymerase, in Arabidopsis thaliana

Mitsumasa Hanaoka, Kengo Kanamaru, Hideo Takahashi and Kan Tanaka*

Laboratory of Molecular Genetics, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan

*To whom correspondence should be addressed. Tel: +81 3 5841 7825; Fax: +81 3 5841 8476; Email: kntanaka{at}iam.u-tokyo.ac.jp

Most photosynthesis-related genes in mature chloroplasts are transcribed by a eubacterial-type RNA polymerase (PEP) whose core subunits are encoded by the plastid genome. It has been shown previously that six putative nuclear genes (SIG1 to SIG6) encode promoter-specificity factors for PEP in Arabidopsis thaliana, and we isolated a T-DNA insertion line of SIG2 (sig2-1 mutant) that manifests aberrant chloroplast development. With the use of S1 nuclease protection and primer extension analyses, we have now characterized the SIG2-dependent chloroplast promoters in A.thaliana. The amounts of transcripts derived from one of the multiple psbD promoters (psbD –256) and from the promoters of two tRNA genes (trnE-UUC and trnV-UAC) were markedly and specifically decreased in the sig2-1 mutant. The abundance of these transcripts was restored to wild-type levels by introduction into the mutant of a SIG2 transgene. The recombinant SIG2 protein mixed with Escherichia coli core RNA polymerase could bind to a DNA fragment that contains the SIG2-dependent psbD –256, trnE-UUC or trnV-UAC promoter. Sequences similar to those of the –35 and –10 promoter elements of E.coli were identified in the regions of the SIG2-dependent chloroplast genes upstream of the transcription initiation sites.


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