Characterization of the Promoter Elements for the Staphylococcal Enterotoxin D Gene

Abstract

Deletion analysis of the promoter for the Staphylococcus aureus enterotoxin D determinant indicated that a 52-bp sequence, from ؊34 to ؉18, was sufficient for sed promoter function and agr regulation. A consensus ؊10 Pribnow box sequence, a less conserved ؊35 sequence, and a TG dinucleotide motif were present. Transcribed sequences (؉1 to ؉18) are essential for promoter activity.Staphylococcal food poisoning is an intoxication resulting from ingestion of foods contaminated with enterotoxin producing Staphylococcus aureus strains (16). The symptoms include emesis, diarrhea, abdominal cramping, and in severe cases, fever and shock (6,30,31). The staphylococcal enterotoxins are a group of secreted proteins that cause emesis when orally administered to primates (6). To date, a number of enterotoxins have been characterized based on their serological reactivities and designated SEA to SEJ, including subtypes SEC 1 to SEC 3 (5,8,24,29,(32)(33)(34).Though staphylococcal enterotoxins are similar in structure and biological properties (22), they differ with respect to genetic localization, amount of toxin produced, and mechanism of gene regulation. The genes for SEA and SEE are carried on prophage, some of which are defective prophage. SED and SEJ gene determinants are carried on the same penicillinasetype plasmid. The genes for SEB and SEC are chromosomal, but the nature of the genetic elements on which they reside has not been elucidated (7,18,19). SEB and SEC are expressed in greater quantities than the other enterotoxins, often on the order of 100 g/ml of culture supernatant, whereas maximal production of SEA, SED, and SEE is usually less than 10 g/ml (3). Furthermore, SEA is produced throughout the log phase of growth, while SEB, SEC, and SED are produced in greater quantities during the transition from the exponential to the stationary phases of growth (5). The latter expression pattern is characteristic of many staphylococcal exoprotein virulence factors which are under the control of the accessory gene regulator (agr) two-component regulatory system (25). The transcription of seb, sec, and sed is subject to regulation by the agr system. In agr mutant strains, mRNA steady-state levels were reduced 4-fold for seb, 5.5-fold for sed, and 2 to 3-fold for sec (2, 28).Information regarding the promoter elements of the staphylococcal enterotoxin genes is very limited. The sea promoter has been identified by means of primer extension analysis in conjunction with deletion mutagenesis. However, detailed characterization of the promoters for the agr-regulated enterotoxin genes has not been reported. Promoter sequences for seb and sed were defined by mapping the transcription start site and by comparison to Escherichia coli promoter consensus Ϫ10 and Ϫ35 sequences. Mahmood and Khan found that a region upstream of the Ϫ35 element, at positions Ϫ93 to Ϫ58, is required for seb expression (21). Although primer extension studies have been carried out on sed, only a Pribnow box could be identified in the sequence (2). No convincing...

Global repression of exotoxin synthesis by staphylococcal superantigens

“…Indeed, the agr-responsive element of sed, which encodes enterotoxin D, has recently been mapped also to within the minimal sed promoter (25). Whether the agr and autorepression targets coincide for seb remains to be determined.…”

Detection of Staphylococcus aureus Enterotoxins A to D by Real-Time Fluorescence PCR Assay

“…We found that the levels of detection of SEB and SEC were genotypically and phenotypically almost identical. Furthermore, SEA is produced throughout the log phase of growth, while SEB, SEC, and SED are produced in greater quantities during the transition from the exponential to the stationary phase of growth (22). However, in accordance with the instructions of the manufacturer, SET-RPLA is performed with overnight liquid cultures, that is, cultures that are in the stationary phase.…”

“…SEA is carried on a prophage (4), and SED is carried by a plasmid (21). The gene for SEC is localized on pathogenicity islands (22). The SEB gene (entB) is also located on a chromosome (13).…”

“…Like the gene for SEE (entE), the gene for SEA (entA) is carried by a temperate bacteriophage (4,6,22). The SEB gene (entB) is located on a chromosome (13).…”

“…The SEB gene (entB) is located on a chromosome (13). The genes for SEC (entC) and TSST-1 are also chromosomal (22). The genes for SED (entD) and SEJ (entJ) are plasmid carried (21).…”