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首页> 美国卫生研究院文献>other >CytR Homolog of Pectobacterium carotovorum subsp. carotovorum Controls Air-Liquid Biofilm Formation by Regulating Multiple Genes Involved in Cellulose Production c-di-GMP Signaling Motility and Type III Secretion System in Response to Nutritional and Environmental Signals
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CytR Homolog of Pectobacterium carotovorum subsp. carotovorum Controls Air-Liquid Biofilm Formation by Regulating Multiple Genes Involved in Cellulose Production c-di-GMP Signaling Motility and Type III Secretion System in Response to Nutritional and Environmental Signals

机译：Carotovorum亚种的CytR同源物。胡萝卜素通过调节涉及营养和环境信号的纤维素生产c-di-GMP信号传导运动性和III型分泌系统中涉及的多个基因来控制气态生物膜的形成

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摘要

Pectobacterium carotovorum subsp. carotovorum [Pcc (formerly Erwinia carotovora subsp. carotovora)] PC1 causes soft-rot disease in a wide variety of plant species by secreting multiple pathogenicity-related traits. In this study, regulatory mechanism of air-liquid (AL) biofilm formation was studied using a cytR homolog gene deletion mutant (ΔcytR) of Pcc PC1. Compared to the wild type (Pcc PC1), the ΔcytR mutant produced fragile and significantly (P < 0.001) lower amounts of AL biofilm on salt-optimized broth plus 2% glycerol (SOBG), yeast peptone dextrose adenine, and also on King’s B at 27°C after 72 h incubation in static condition. The wild type also produced significantly higher quantities of AL biofilm on SOBGMg– (magnesium deprived) containing Cupper (Cu2+), Zinc (Zn2+), Manganese (Mn2+), Magnesium (Mg2+), and Calcium (Ca2+) compared to the ΔcytR mutant. Moreover, the wild type was produced higher amounts of biofilms compared to the mutant while responding to pH and osmotic stresses. The ΔfliC (encoding flagellin), flhD::Tn5 (encoding a master regulator) and ΔmotA (a membrane protein essential for flagellar rotation) mutants produced a lighter and more fragile AL biofilm on SOBG compared to their wild counterpart. All these mutants resulted in having weak bonds with the cellulose specific dye (Calcofluor) producing lower quantities of cellulose compared to the wild type. Gene expression analysis using mRNA collected from the AL biofilms showed that ΔcytR mutant significantly (P < 0.001) reduced the expressions of multiple genes responsible for cellulose production (bcsA, bcsE, and adrA), motility (flhD, fliA, fliC, and motA) and type III secretion system (hrpX, hrpL, hrpA, and hrpN) compared to the wild type. The CytR homolog was therefore, argued to be able to regulate the AL biofilm formation by controlling cellulose production, motility and T3SS in Pcc PC1. In addition, all the mutants exhibited poorer attachment to radish sprouts and AL biofilm cells of the wild type was resistant than stationary-phase and planktonic cells to acidity and oxidative stress compared to the same cells of the ΔcytR mutant. The results of this study therefore suggest that CytR homolog is a major determinant of Pcc PC1’s virulence, attachment and its survival mechanism.
机译：胡萝卜杆菌假单胞菌亚种。 carotovorum [Pcc（以前的欧文氏carotovora亚种carotovora）] PC1通过分泌多种致病性相关性状，在多种植物中引起软腐病。在这项研究中，使用Pcc PC1的cytR同源基因缺失突变体（ΔcytR）研究了气液（AL）生物膜形成的调控机制。与野生型（Pcc PC1）相比，ΔcytR突变体在盐优化的肉汤加2％甘油（SOBG），酵母蛋白one右旋糖腺嘌呤和King's B上产生的脆弱且显着降低（P <0.001）的AL生物膜量在静态条件下孵育72小时后，在27°C下保存。野生型还在含有铜（Cu 2 + ），锌（Zn 2 + – （缺镁）上产生大量的AL生物膜。 sup>），锰（Mn 2 + ），镁（Mg 2 + ）和钙（Ca 2 + ）与ΔcytR相比突变体。此外，与突变体相比，野生型产生了更高数量的生物膜，同时对pH和渗透压产生了响应。 ΔfliC（编码鞭毛蛋白）， flhD :: Tn5（编码主调节剂）和Δ motA （鞭毛旋转必不可少的膜蛋白）突变体产生的重量更轻，更脆弱与野生动物相比，SOBG上的AL生物膜。与野生型相比，所有这些突变体均导致与纤维素特定染料（Calcofluor）的键弱，产生的纤维素量较少。使用从AL生物膜收集的mRNA进行的基因表达分析表明，Δ cytR 突变体显着（ P <0.001）降低了负责纤维素生产的多个基因（ bcsA）的表达。 ，bcsE 和 adrA ），运动性（ flhD，fliA，fliC 和 motA ）和III型分泌系统（< em> hrpX，hrpL，hrpA 和 hrpN ）。因此，据称CytR同源物能够通过控制Pcc PC1中的纤维素产生，运动性和T3SS来调节AL生物膜的形成。此外，与Δ cytR 的相同细胞相比，所有突变体对萝卜芽的附着力均较差，野生型的AL生物膜细胞对酸性和氧化应激的抵抗力高于固定相和浮游细胞。突变体。因此，这项研究的结果表明CytR同源物是Pcc PC1的毒力，附着力及其生存机制的主要决定因素。

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