title: Eukaryotic-type plastid nucleoid protein pTAC3 is essential for transcription by the bacterial-type plastid RNA polymerase.
abstract: Plastid transcription is mediated by two distinct types of RNA polymerases (RNAPs), bacterial-type RNAP (PEP) and phage-type RNAP (NEP). Recent genomic and proteomic studies revealed that higher plants have lost most prokaryotic transcription regulators and have acquired eukaryotic-type proteins during plant evolution. However, in vivo dynamics of chloroplast RNA polymerases and eukaryotic-type plastid nucleoid proteins have not been directly characterized experimentally. Here, we examine the association of the α-subunit of PEP and eukaryotic-type protein, plastid transcriptionally active chromosome 3 (pTAC3) with transcribed regions in vivo by using chloroplast chromatin immunoprecipitation (cpChIP) assays. PEP α-subunit preferentially associates with PEP promoters of photosynthesis and rRNA genes, but not with NEP promoter regions, suggesting selective and accurate recognition of PEP promoters by PEP. The cpChIP assays further demonstrate that the peak of PEP association occurs at the promoter-proximal region and declines gradually along the transcribed region. pTAC3 is a putative DNA-binding protein that is localized to chloroplast nucleoids and is essential for PEP-dependent transcription. Density gradient and immunoprecipitation analyses of PEP revealed that pTAC3 is associated with the PEP complex. Interestingly, pTAC3 associates with the PEP complex not only during transcription initiation, but also during elongation and termination. These results suggest that pTAC3 is an essential component of the chloroplast PEP complex. In addition, we demonstrate that light-dependent chloroplast transcription is mediated by light-induced association of the PEP-pTAC3 complex with promoters. This study illustrates unique dynamics of PEP and its associated protein pTAC3 during light-dependent transcription in chloroplasts.
authors.affiliation: Faculty of Agriculture and Institute of Advanced Study,  Kyushu University, Fukuoka 812-8581,  Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Yagi
authors.forename: Yoko
authors.initials: Y
authors.lastname: Ishizaki
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Nakahira
authors.forename: Yuzuru
authors.initials: Y
authors.lastname: Tozawa
authors.forename: Takashi
authors.initials: T
authors.lastname: Shiina
mesh_terms: D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D001419:Bacteria / Q000201:enzymology; D060365:Chloroplast Proteins / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D047369:Chromatin Immunoprecipitation; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant / Q000528:radiation effects; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D015151:Immunoblotting; D008027:Light; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding / Q000528:radiation effects; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011523:Protoplasts / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic
chemical_list: D029681:Arabidopsis Proteins; D060365:Chloroplast Proteins; D021122:Protein Subunits; D049452:Green Fluorescent Proteins; D012321:DNA-Directed RNA Polymerases; C113275:RNA polymerase alpha subunit