ABSTRACT

Objective Treponema denticola is involved in ‘chronic’ periodontitis pathogenesis. The mechanism underlying the regulation of the expression of its virulence factors, such as major surface protein (Msp) and prolyl-phenylalanine specific protease (dentilisin) is yet to be clarified. We determined the gene expression profiles of Msp- and dentilisin-deficient mutants of T. denticola to identify the regulation network of gene expression concomitant with the inactivation of these virulence genes.

Methods Gene expression profiles of T. denticola ATCC 35405 (wild type), dentilisin-deficient mutant K1, and msp-deficient mutant DMSP3 were determined using DNA microarray analysis and quantitative real-time reverse transcription PCR (qRT-PCR). Msp and dentilisin protein levels were determined by immunoblotting and proteolytic activity assays.

Results In addition to several differentially expressed genes, dentilisin expression was reduced in DMSP3; msp expression was significantly reduced in K1 (p < 0.05), both at the gene and protein levels. To identify the regulatory system involved, the expression levels of the potential regulators whose expression showed changes in the mutants were evaluated using qRT-PCR. Transcriptional regulators TDE_0127 and TDE_0814 were upregulated in K1, and the potential repressor, TDE_0344, was elevated in DMSP3.

Conclusions Dentilisin and Msp expression were interrelated, and gene expression regulators, such as TDE_0127, may be involved in their regulation.

摘要

目的 齿状螺旋体参与“慢性”牙周炎的发病机制。其毒力因子，如主要表面蛋白 (Msp) 和脯氨酰苯丙氨酸特异性蛋白酶 (dentilisin) 表达的调控机制尚未阐明。我们确定了T. denticola的 Msp 和牙菌素缺陷突变体的基因表达谱，以确定伴随这些毒力基因失活的基因表达调控网络。

方法的基因表达谱T.齿垢ATCC 35405（野生型），dentilisin缺陷型突变K1，和MSP-使用DNA微阵列分析和定量实时逆转录PCR（QRT-PCR）测定缺陷突变DMSP3。Msp 和牙菌素蛋白水平通过免疫印迹和蛋白水解活性测定确定。

结果除了几个差异表达的基因外，DMSP3 中牙菌素的表达降低；msp表达在 K1 中显着降低 (p < 0.05)，在基因和蛋白质水平上。为了确定所涉及的调控系统，使用 qRT-PCR 评估了在突变体中表达发生变化的潜在调控因子的表达水平。转录调节因子 TDE_0127 和 TDE_0814 在 K1 中上调，潜在抑制因子 TDE_0344 在 DMSP3 中升高。

结论Dentilisin与Msp的表达存在相关性，TDE_0127等基因表达调控因子可能参与了它们的调控。