Molecular genetic analysis of chloroplast gene promoters dependent on SIG2, a nucleus-encoded sigma factor for the plastid-encoded RNA polymerase, in Arabidopsis thaliana.
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.
Hanaoka
Mitsumasa
M
Laboratory of Molecular Genetics, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan.
Kanamaru
Kengo
K
Takahashi
Hideo
H
Tanaka
Kan
K
D017360:Arabidopsis / Q000166:cytology / Q000201:enzymology / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics; D001483:Base Sequence; D002467:Cell Nucleus / Q000235:genetics*; D002736:Chloroplasts / Q000235:genetics*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D017343:Genes, Plant / Q000235:genetics; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D023282:Nuclease Protection Assays; D018087:Plastids / Q000201:enzymology*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D024363:Transcription Initiation Site; D014158:Transcription, Genetic / Q000235:genetics; D019076:Transgenes / Q000235:genetics
D029681:Arabidopsis Proteins; D004268:DNA-Binding Proteins; D018749:RNA, Plant; C413368:Sig2 sigma factor; D012808:Sigma Factor; D004247:DNA; D012343:RNA, Transfer; D012321:DNA-Directed RNA Polymerases