Ostrowski J, Kredich N M
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
J Bacteriol. 1991 Apr;173(7):2212-8. doi: 10.1128/jb.173.7.2212-2218.1991.
CysB protein positively regulates genes of the Salmonella typhimurium cysteine regulon and negatively autoregulates cysB. The cysB promoter was characterized by primer extension of cellular RNA, which gave products identifying a major in vivo transcription start site located 95 bp upstream of the cysB start codon and two minor sites located 9 and 10 bp downstream of the major site. Gel shift binding studies and DNase I footprinting experiments showed that CysB protein binds to the cysB promoter from position -10 to +36 relative to the major transcription start site. We have designated this binding site CBS-B. CysB protein inhibited transcription initiation at the cysB promoter in an in vitro runoff assay, indicating that cysB is negatively autoregulated by the binding of CysB protein to the cysB promoter, where it acts as a repressor. N-Acetyl-L-serine, an inducer of the cysteine regulon, inhibited the binding of CysB protein to the cysB promoter and partially reversed the ability of CysB protein to inhibit transcription initiation. These effects are in contrast to those observed in studies of positively regulated cys promoters, in which N-acetyl-L-serine stimulates binding and causes CysB protein to activate transcription initiation.
CysB蛋白正向调控鼠伤寒沙门氏菌半胱氨酸调节子的基因，并负向自调控cysB。通过对细胞RNA进行引物延伸来表征cysB启动子，其产物确定了一个主要的体内转录起始位点，位于cysB起始密码子上游95 bp处，以及两个次要位点，位于主要位点下游9和10 bp处。凝胶迁移结合研究和DNase I足迹实验表明，CysB蛋白与cysB启动子结合，相对于主要转录起始位点，结合位置为-10至+36。我们将这个结合位点命名为CBS-B。在体外径流分析中，CysB蛋白抑制了cysB启动子处的转录起始，这表明cysB通过CysB蛋白与cysB启动子的结合而受到负向自调控，在该启动子处CysB蛋白作为阻遏物发挥作用。半胱氨酸调节子的诱导剂N-乙酰-L-丝氨酸抑制了CysB蛋白与cysB启动子的结合，并部分逆转了CysB蛋白抑制转录起始的能力。这些效应与在正向调控的cys启动子研究中观察到的效应相反，在正向调控的cys启动子研究中，N-乙酰-L-丝氨酸刺激结合并导致CysB蛋白激活转录起始。
