Fraser Gillian M, Claret Laurent, Furness Richard, Gupta Srishti, Hughes Colin
Cambridge University Department of Pathology, Tennis Court Road, Cambridge CB2 1QP, UK1.
Microbiology (Reading). 2002 Jul;148(Pt 7):2191-2201. doi: 10.1099/00221287-148-7-2191.
The HpmA haemolysin toxin of Proteus mirabilis is encoded by the hpmBA locus and its production is upregulated co-ordinately with the synthesis and assembly of flagella during differentiation into hyperflagellated swarm cells. Primer extension identified a sigma(70) promoter upstream of hpmB that was upregulated during swarming. Northern blotting indicated that this promoter region was also required for concomitant transcription of the immediately distal hpmA gene, and that the unstable hpmBA transcript generated a stable hpmA mRNA and an unstable hpmB mRNA. Transcriptional luxAB fusions to the DNA regions 5' of the hpmB and hpmA genes confirmed that hpmB sigma(70) promoter activity increased in swarm cells, and that there was no independent hpmA promoter. Increased transcription of the hpmBA operon in swarm cells was dependent upon a 125 bp sequence 5' of the sigma(70) promoter -35 hexamer. This sequence spans multiple putative binding sites for the leucine-responsive regulatory protein (Lrp), and band-shift assays with purified Lrp confirmed the presence of at least two such sites. The influence on hpmBA expression of the key swarming positive regulators FlhD(2)C(2) (encoded by the flagellar master operon), Lrp, and the membrane-located upregulator of the master operon, UmoB, was examined. Overexpression of each of these regulators moderately increased hpmBA transcription in wild-type P. mirabilis, and the hpmBA operon was not expressed in any of the flhDC, lrp or umoB mutants. Expression in the mutants was not recovered by cross-complementation, i.e. by overexpression of FlhD(2)C(2), Lrp or UmoB. Expression of the zapA protease virulence gene, which like hpmBA is also upregulated in swarm cells, did not require Lrp, but like flhDC it was upregulated by UmoB. The results indicate intersecting pathways of control linking virulence gene expression and swarm cell differentiation.
奇异变形杆菌的HpmA溶血素毒素由hpmBA基因座编码，在分化为超鞭毛群集细胞的过程中，其产生与鞭毛的合成和组装协同上调。引物延伸鉴定出hpmB上游的一个σ⁷⁰启动子，该启动子在群集过程中上调。Northern印迹表明，该启动子区域对于紧邻其下游的hpmA基因的伴随转录也是必需的，并且不稳定的hpmBA转录本产生了稳定的hpmA mRNA和不稳定的hpmB mRNA。与hpmB和hpmA基因5'端DNA区域的转录luxAB融合证实，hpmB σ⁷⁰启动子活性在群集细胞中增加，并且不存在独立的hpmA启动子。群集细胞中hpmBA操纵子转录的增加依赖于σ⁷⁰启动子-35六聚体5'端的一个125 bp序列。该序列跨越多个推测的亮氨酸响应调节蛋白（Lrp）结合位点，用纯化的Lrp进行的凝胶迁移试验证实至少存在两个这样的位点。研究了关键群集正调节因子FlhD₂C₂（由鞭毛主操纵子编码）、Lrp和主操纵子的膜定位上调因子UmoB对hpmBA表达的影响。这些调节因子中的每一个的过表达都适度增加了野生型奇异变形杆菌中hpmBA的转录，并且hpmBA操纵子在任何flhDC、lrp或umoB突变体中均不表达。突变体中的表达不能通过交叉互补恢复，即通过过表达FlhD₂C₂、Lrp或UmoB来恢复。zapA蛋白酶毒力基因的表达，其与hpmBA一样在群集细胞中也上调，不需要Lrp，但与flhDC一样，它受UmoB上调。结果表明，连接毒力基因表达和群集细胞分化的控制途径相互交叉。
