Involvement of CysB and Cbl regulatory proteins in expression of the tauABCD operon and other sulfate starvation-inducible genes in Escherichia coli

Abstract

Starvation for sulfate results in increased synthesis of several proteins in Escherichia coli. Among these Ssi (sulfate starvation-induced) proteins are the products of the tauABCD genes, which are required for utilization of taurine as sulfur source for growth. In this study, the role of the cbl gene in expression of tauABCD and other ssi genes was investigated. The protein encoded by cbl shows high sequence similarity to CysB, the LysR-type transcriptional activator of the genes involved in cysteine biosynthesis. Strain EC2541, which contains an internal deletion in cbl, was unable to utilize taurine and other aliphatic sulfonates as sulfur sources. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that many of the Ssi proteins were not synthesized in EC2541. Expression of a translational tauD-lacZ fusion required the presence of both cbl and cysB. The interactions of CysB and Cbl with the promoter region of tauABCD were studied by using gel mobility shift experiments and DNase I footprinting. CysB occupied multiple binding sites, whereas Cbl occupied only one site from 112 to 68 bp upstream of the transcription start site. Acetylserine, the inducer of transcription of CysB-regulated genes, stimulated binding of CysB but not of Cbl. Sulfate had no effect on binding of both proteins to the tauABCD promoter region. These results indicate that Cbl is a transcription factor for genes required for sulfonate-sulfur utilization and maybe for other genes whose expression is induced by sulfate starvation.Cysteine biosynthesis via sulfate assimilation in plants and microorganisms represents the predominant mechanism by which inorganic sulfur is reduced and incorporated into organic compounds. This pathway has been characterized in detail in Escherichia coli and Salmonella typhimurium (see reference 24 for a recent review). Most of the genes involved are coordinately regulated as the cysteine regulon-high-level expression of these genes requires sulfur limitation, the LysRtype (11, 34) transcriptional regulator CysB, and the presence of the inducer molecule N-acetyl-L-serine. Transcription from cys gene promoters is initiated by CysB protein binding to activation sites positioned just upstream of the Ϫ35 regions of these promoters, and inducer facilitates interaction of CysB with these sites (15,23,28). Sulfur limitation is a necessary condition for derepression of the cys regulon because L-cysteine inhibits the serine transacetylase enzyme required for inducer synthesis. In addition, sulfide and thiosulfate act as anti-inducers of CysB-dependent transcription (13, 23). Maximal expression of cys genes is seen during growth with limiting sulfur sources such as glutathione or L-djenkolic acid, whereas growth with sulfate, sulfite, or thiosulfate leads to partial repression of these genes, and they are fully repressed during growth with sulfide, L-cysteine, and L-cystine (22,24). The partial repression of cys gene expression by sulfate is ascribed to the inhibitory effect of sulfide generated ...

Functional Dissection of the LysR-type CysB Transcriptional Regulator

“…Examples of LysR family members, NahR (7), OxyR (9), and GcvA (36), also suggest the correlation between the oligomerization defect and the inability of the mutant protein to exert a dominant-negative effect (also called a "poisoning effect") on the activity of the wild-type counterpart produced by the cell. For CysB, the negative dominance was clearly seen with some non-repressing (non-binding) variants (with mutations in region [11][12][13][14][15][16][17][18][19][20][21][22] as well as with non-inducible variants (Fig. 1).…”

Nitrogen regulatory protein C-controlled genes of Escherichia coli : Scavenging as a defense against nitrogen limitation

“…The difference in mRNA levels-up to 30-fold higher in the gln-L(Up) strain-was one of the largest we observed. Although we could not demonstrate a clear physiological role for Cbl in nitrogen metabolism, it activates transcription of operons whose products catabolize nitrogen-and sulfur-containing compounds (51). The cbl mRNA in glnL(Up) strains appeared to be full length (not shown), and we hope to determine whether it is translated to yield protein.…”

A CysB-regulated and σ54-dependent regulator, SfnR, is essential for dimethyl sulfone metabolism of Pseudomonas putida strain DS1

“…Several enteric and soil bacteria are able to utilize a variety of organosulfur compounds including taurine, MSA, alkanesulfonates and/or alkanesulfate esters as sulfur sources (Roberts et al, 1955;Uria-Nickelsen et al, 1993;van der Ploeg et al, 1996van der Ploeg et al, , 1998van der Ploeg et al, , 1999Hummerjohann et al, 1998;Vermeij et al, 1999;Kertesz, 1999). Recently, the genes involved in utilization of such organosulfur compounds in Escherichia coli, Bacillus subtilis, Pseudomonas putida and P. aeruginosa have been reported.…”

“…Recently, the genes involved in utilization of such organosulfur compounds in Escherichia coli, Bacillus subtilis, Pseudomonas putida and P. aeruginosa have been reported. The tauABCD and ssuEADCB operons of E. coli are required for taurine and alkanesulfonate utilization, respectively (van der Ploeg et al, 1996(van der Ploeg et al, , 1999. The expression of tau and ssu operons is repressed in the presence of preferred sulfur sources such as inorganic sulfate, thiosulfate or cysteine, but induced in their absence (van der Ploeg et al, 1997(van der Ploeg et al, , 1999.…”

“…The tauABCD and ssuEADCB operons of E. coli are required for taurine and alkanesulfonate utilization, respectively (van der Ploeg et al, 1996(van der Ploeg et al, , 1999. The expression of tau and ssu operons is repressed in the presence of preferred sulfur sources such as inorganic sulfate, thiosulfate or cysteine, but induced in their absence (van der Ploeg et al, 1997(van der Ploeg et al, , 1999. This phenomenon is called the sulfate starvation response (SSR; Quadroni et al, 1996), and is controlled by two LysR-type transcriptional regulators, CysB and Cbl, in E. coli (van der Ploeg et al, 1997(van der Ploeg et al, , 1999Bykowski et al, 2002).…”