Interconnections between Sigma B, agr, and Proteolytic Activity in Staphylococcus aureus Biofilm Maturation
Interconnections between Sigma B, agr, and Proteolytic Activity in Staphylococcus aureus Biofilm Maturation
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DOI：
10.1128/iai.01036-08
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发表时间：
2009-04-01
影响因子：
3.1
通讯作者：
Horswill, Alexander R.
中科院分区：
文献类型：
--
作者：
Lauderdale, Katherine J.;Boles, Blaise R.;Horswill, Alexander R.
金黄色葡萄球菌是宿主组织和医疗植入物上的熟练生物膜形成者。我们对S.金黄色葡萄球菌菌株SH 1000中鉴定生物膜成熟的ICA非依赖性机制所必需的基因座，并鉴定rsbUVW-sigB操纵子中的多个插入。在构建和表征sigB缺失后，我们确定生物膜表型是由于缺乏sigma因子B（SigB）活性。该表型在USA 300菌株LAC的sigB突变体中是保守的，该菌株LAC是一种被充分研究的社区相关的耐甲氧西林S。金黄色葡萄球菌分离株我们确定sigB突变体中agr RNAIII水平升高，并且已知高水平的RNAIII表达具有膜效应。通过将agr突变引入SH 1000或LAC sigB缺失菌株，S.金黄色葡萄球菌恢复了生物膜能力，表明生物膜表型是AGR依赖性的。蛋白酶活性与agr活性和ica独立的生物膜形成有关，我们观察到蛋白酶抑制剂苯甲基磺酰氟和α-巨球蛋白可以逆转sigB生物膜缺陷。类似地，在sigB突变体中失活编码金霉素和Spl细胞外蛋白酶的基因恢复了生物膜能力。由于胞壁蛋白水解酶活性和生物膜成熟之间的联系越来越大，进行自溶素酶谱分析，结果显示sigB突变体的谱发生了改变;再次，表型可以通过蛋白酶失活修复。这些发现表明，SigB活性的缺乏导致RNAIII表达增加，从而提高细胞外蛋白酶水平并改变胞壁蛋白水解酶活性谱。总之，我们的观察表明SigB是S的重要调节因子。金黄色葡萄球菌生物膜成熟。
Staphylococcus aureus is a proficient biofilm former on host tissues and medical implants. We mutagenized S. aureus strain SH1000 to identify loci essential for ica-independent mechanisms of biofilm maturation and identified multiple insertions in the rsbUVW-sigB operon. Following construction and characterization of a sigB deletion, we determined that the biofilm phenotype was due to a lack of sigma factor B (SigB) activity. The phenotype was conserved in a sigB mutant of USA300 strain LAC, a well-studied community-associated methicillin-resistant S. aureus isolate. We determined that agr RNAIII levels were elevated in the sigB mutants, and high levels of RNAIII expression are known to have antibiofilm effects. By introducing an agr mutation into the SH1000 or LAC sigB deletion strain, S. aureus regained biofilm capacity, indicating that the biofilm phenotype was agr dependent. Protease activity is linked to agr activity and ica-independent biofilm formation, and we observed that the protease inhibitors phenylmethylsulfonyl fluoride and alpha-macroglobulin could reverse the sigB biofilm defect. Similarly, inactivating genes encoding both the aureolysin and Spl extracellular proteases in the sigB mutant restored biofilm capacity. Due to the growing link between murein hydrolase activity and biofilm maturation, autolysin zymography was performed, which revealed an altered profile in the sigB mutant; again, the phenotype could be repaired through protease inactivation. These findings indicate that the lack of SigB activity results in increased RNAIII expression, thus elevating extracellular protease levels and altering the murein hydrolase activity profile. Altogether, our observations demonstrate that SigB is an essential regulator of S. aureus biofilm maturation.