Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, United States.
Department of Chemistry and Biochemistry, Brigham Young University, Provo, UT, United States.
Front Cell Infect Microbiol. 2018 Sep 18;8:323. doi: 10.3389/fcimb.2018.00323. eCollection 2018.
The formation of biofilms by () and requires the genes, which direct production of a polysaccharide extracellular matrix (Hms-ECM). Despite possessing identical sequences, produces much less Hms-ECM than . The regulatory influences that control Hms-ECM production and biofilm formation are not fully understood. In this study, negative regulators of biofilm production in were identified. Inactivation of the BarA/UvrY two-component system or the CsrB regulatory RNA increased binding of Congo Red dye, which correlates with extracellular polysaccharide production. These mutants also produced biofilms that were substantially more cohesive than the wild type strain. Disruption of was not sufficient for to cause proventricular blockage during infection of fleas. However, this strain was less acutely toxic toward fleas than wild type . Flow cytometry measurements of lectin binding indicated that BarA/UvrY/CsrB mutants may produce higher levels of other carbohydrates in addition to poly-GlcNAc Hms-ECM. In an effort to characterize the relevant downstream targets of the BarA/UvrY system, we conducted a proteomic analysis to identify proteins with lower abundance in the ::Tn5 mutant strain. Urease subunit proteins were less abundant and urease enzymatic activity was lower, which likely reduced toxicity toward fleas. Loss of CsrB impacted expression of several potential regulatory proteins that may influence biofilms, including the RcsB regulator. Overexpression of CsrB did not alter the Congo-red binding phenotype of an ::Tn5 mutant, suggesting that the effect of CsrB on biofilms may require RcsB. These results underscore the regulatory and compositional differences between and biofilms. By activating CsrB expression, the BarA/UvrY two-component system has pleiotropic effects that impact biofilm production and stability.
生物膜的形成需要 () 和 基因，这些基因指导多糖细胞外基质 (Hms-ECM) 的产生。尽管具有相同的 序列，但 产生的 Hms-ECM 比 少得多。控制 生产和生物膜形成的调节影响尚未完全理解。在这项研究中，鉴定了 生物膜形成的负调节因子。BarA/UvrY 双组分系统或 CsrB 调节 RNA 的失活增加了刚果红染料的结合，这与细胞外多糖的产生相关。这些突变体还产生了比野生型菌株更具凝聚力的生物膜。 的破坏不足以在感染跳蚤时导致前胃阻塞。然而，这种菌株对跳蚤的急性毒性比野生型 低。凝集素结合的流式细胞术测量表明， BarA/UvrY/CsrB 突变体除了聚 GlcNAc Hms-ECM 之外，可能还产生其他碳水化合物的水平更高。为了表征 BarA/UvrY 系统的相关下游靶标，我们进行了蛋白质组学分析，以鉴定在 ::Tn5 突变株中丰度较低的蛋白质。脲酶亚基蛋白丰度较低，脲酶酶活性较低，这可能降低了对跳蚤的毒性。CsrB 的缺失影响了几个潜在调节蛋白的表达，这些蛋白可能影响生物膜，包括 RcsB 调节剂。CsrB 的过表达并未改变 ::Tn5 突变体的刚果红结合表型，这表明 CsrB 对生物膜的影响可能需要 RcsB。这些结果强调了 和 生物膜之间的调节和组成差异。通过激活 CsrB 的表达， BarA/UvrY 双组分系统具有影响生物膜产生和稳定性的多效性效应。
