Curation Information

Publication: GacA-controlled activation of promoters for small RNA genes in Pseudomonas fluorescens.;Humair B, Wackwitz B, Haas D;Applied and environmental microbiology 2010 Mar; 76(5):1497-506 [20048056]
TF: PsrA [Q4KFB4, view regulon]
Reported TF sp.: Pseudomonas fluorescens CHA0
Reported site sp.: Pseudomonas fluorescens CHA0
Created by: Dinara Sagitova
Curation notes: -

Experimental Process

Multiple sequence alignment of rsmX, rsmY, and rsmZ promoters revealed a conserved palindromic UAS. The rsmZ-lacZ reporter assays showed that deletion of the UAS in rsmZ promoter resulted in almost complete loss of transcription in strain CHA0 growing in NYB. The rsmZ-lacZ fusion showed reduced and delayed expression in the psrA deletion mutant compared to that in wild-type strain. A potential PsrA recognition sequence was found overlapping with the UAS in the rsmZ promoter region. When this sequence was mutated either by a 3-bp deletion or by a 3-bp change, neither of which altered the UAS, regulation by PsrA appeared to be lost.

Transcription Factor Binding Sites

Sites reported in the paper
Sites matched in the genome

CAAAGATTACTTTT

Gene Regulation

Regulated genes for each binding site are displayed below. Gene regulation diagrams show binding sites, positively-regulated genes, negatively-regulated genes, both positively and negatively regulated genes, genes with unspecified type of regulation. For each indvidual site, experimental techniques used to determine the site are also given.

Site sequence	Regulated genes	Gene diagram	Experimental techniques	TF function	TF type
CAAAGATTACTTTT	rsmZ		Experimental technique details Beta-gal reporter assay Beta-gal reporter assay Reporter assay using the beta-galactosidase (lacZ) gene. The lacZ gene is typically fused to the promoter of interest. Differential regulation of the promoter mediated by the TF is assessed by induction of the system and evaluation of lacZ expression. Bacteria expressing lacZ appear blue when grown on a X-gal medium. The assay is often performed using a plasmid borne construction on a lacZ(def) strain. - Experimental technique details Multiple sequence alignment (MSA) (ECO:0005556) Multiple sequence alignment (MSA) - ECO:0005556 MSA can be used to align promoter regions and highlight regions of conservation that may be indicative of transcription factor binding sites. - Experimental technique details Site directed mutagenesis (ECO:0005667) Site directed mutagenesis - ECO:0005667 Target-specific mutation, as opposed to non-specific mutation. In the context of TF-binding sites, site-directed mutagenesis is typically used to establish/confirm the specific sequence and location of a site, often in tandem with EMSA. Different positions of a putative binding site are mutated to non-consensus (or random) bases and binding to the mutated site is evaluated through EMSA or other means. Often implemented only in conserved motif positions or serially through all positions of a site. - Experimental technique details Visual sequence inspection (nan) Visual sequence inspection - nan Visual inspection of promoter region to identify putative binding sites based on previous knowledge -	activator	not specified