vimA gene downstream of recA is involved in virulence modulation in Porphyromonas gingivalis W83
vimA gene downstream of recA is involved in virulence modulation in Porphyromonas gingivalis W83
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DOI：
10.1128/iai.69.1.325-335.2001
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发表时间：
2001-01-01
影响因子：
3.1
通讯作者：
Fletcher, HM
中科院分区：
文献类型：
--
作者：
Abaibou, H;Chen, Z;Fletcher, HM
在牙龈卟啉单胞菌的recA基因下游鉴定出编码独特的32-kDa蛋白质的0.9-kb开放阅读框。逆转录PCR和Northern印迹分析表明recA基因和该开放阅读框是同一转录单位的一部分。使用 ermF-ermAM 抗生素抗性盒将该克隆片段插入失活，通过等位基因交换产生有缺陷的突变体。当铺在布鲁氏菌血琼脂上时，命名为牙龈卟啉单胞菌 FLL92 的突变菌株呈非黑色，与亲本菌株牙龈卟啉单胞菌 W83 相比，β-溶血显着减少。精氨酸和赖氨酸特异性半胱氨酸蛋白酶活性大部分为可溶性，比亲本菌株低约 90%。 rgpA、rgpB 和 kgp 蛋白酶基因的表达在牙龈卟啉单胞菌 FLL92 中与野生型菌株中相同。与亲本菌株相比，牙龈卟啉单胞菌FLL92除了血凝和溶血素活性显着降低外，还表现出自聚集增加。在使用小鼠模型的体内实验中，牙龈卟啉单胞菌 FLL92 的毒性明显低于亲本菌株。使用所有已知的牙龈卟啉单胞菌插入序列元件作为探针对该突变体和亲本菌株进行的分子调查表明，牙龈卟啉单胞菌FLL92中没有发生由于这些元件的移动而导致的基因组内变化。综上所述，这些结果表明，recA下游基因，即vimA（毒力调节基因），在牙龈卟啉单胞菌W83的毒力调节中发挥着重要作用，可能代表牙龈卟啉单胞菌毒力因子的一种新的转录后或翻译调节。
A 0.9-kb open reading frame encoding a unique 32-kDa protein was identified downstream of the recA gene of Porphyromonas gingivalis. Reverse transcription PCR and Northern blot analysis showed that both the recA gene and this open reading frame are part of the same transcriptional unit. This cloned fragment was insertionally inactivated using the ermF-ermAM antibiotic resistance cassette to create a defective mutant by allelic exchange. When plated on Brucella blood agar, the mutant strain, designated P. gingivalis FLL92, was non-black pigmented and showed significant reduction in beta-hemolysis compared with the parent strain, P. gingivalis W83. Arginine- and lysine-specific cysteine protease activities, which were mostly soluble, were approximately 90% lower than that of the parent strain. Expression of the rgpA, rgpB, and kgp protease genes was the same in P. gingivalis FLL92 as in the wild-type strain. In contrast to the parent strain, P. gingivalis FLL92 showed increased autoaggregration in addition to a significant reduction in hemagglutinating and hemolysin activities. In in vivo experiments using a mouse model, P. gingivalis FLL92 was dramatically less virulent than the parent strain. A molecular survey of this mutant and the parent strain using all known P. gingivalis insertion sequence elements as probes suggested that no intragenomic changes due to the movement of these elements have occurred in P. gingivalis FLL92. Taken together, these results suggest that the recA downstream gene, designated vimA (virulence-modulating gene), plays an important role in virulence modulation in P. gingivalis W83, possibly representing a novel posttranscriptional or translational regulation of virulence factors in P. gingivalis.