title: Bacillus subtilis δ Factor Functions as a Transcriptional Regulator by Facilitating the Open Complex Formation.
abstract: Most bacterial RNA polymerases (RNAP) contain five conserved subunits, viz. 2α, β, β', and ω. However, in many Gram-positive bacteria, especially in fermicutes, RNAP is associated with an additional factor, called δ. For over three decades since its identification, it had been thought that δ functioned as a subunit of RNAP to enhance the level of transcripts by recycling RNAP. In support of the previous observations, we also find that δ is involved in recycling of RNAP by releasing the RNA from the ternary complex. We further show that δ binds to RNA and is able to recycle RNAP when the length of the nascent RNA reaches a critical length. However, in this work we decipher a new function of δ. Performing biochemical and mutational analysis, we show that Bacillus subtilis δ binds to DNA immediately upstream of the promoter element at A-rich sequences on the abrB and rrnB1 promoters and facilitates open complex formation. As a result, δ facilitates RNAP to initiate transcription in the second scale, compared with minute scale in the absence of δ. Using transcription assay, we show that δ-mediated recycling of RNAP cannot be the sole reason for the enhancement of transcript yield. Our observation that δ does not bind to RNAP holo enzyme but is required to bind to DNA upstream of the -35 promoter element for transcription activation suggests that δ functions as a transcriptional regulator.
authors.affiliation: From the Department of Chemistry, Bose Institute, Kolkata-700009, India.
authors.forename: Ranjit Kumar
authors.initials: RK
authors.lastname: Prajapati
authors.affiliation: From the Department of Chemistry, Bose Institute, Kolkata-700009, India.
authors.forename: Shreya
authors.initials: S
authors.lastname: Sengupta
authors.affiliation: From the Department of Chemistry, Bose Institute, Kolkata-700009, India.
authors.forename: Paulami
authors.initials: P
authors.lastname: Rudra
authors.affiliation: From the Department of Chemistry, Bose Institute, Kolkata-700009, India jayanta@jcbose.ac.in.
authors.forename: Jayanta
authors.initials: J
authors.lastname: Mukhopadhyay
mesh_terms: D020856:AT Rich Sequence; D001412:Bacillus subtilis / Q000502:physiology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D015964:Gene Expression Regulation, Bacterial; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D012336:RNA, Ribosomal, 16S / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012235:Rhodamines / Q000737:chemistry; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D024363:Transcription Initiation Site; D061785:Transcription Initiation, Genetic; D014158:Transcription, Genetic
chemical_list: C060762:AbrB protein, Bacillus subtilis; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; D012336:RNA, Ribosomal, 16S; D011994:Recombinant Proteins; D012235:Rhodamines; D012808:Sigma Factor; D014157:Transcription Factors; C005358:tetramethylrhodamine; D012321:DNA-Directed RNA Polymerases
keywords: Bacillus subtilis; DNA binding protein; RNA polymerase; delta factor; gram-positive bacteria; transcription; transcription regulation; transcriptional regulator