A Genome-Wide Analysis of Adhesion in Caulobacter crescentus Identifies New Regulatory and Biosynthetic Components for Holdfast Assembly
A Genome-Wide Analysis of Adhesion in Caulobacter crescentus Identifies New Regulatory and Biosynthetic Components for Holdfast Assembly
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DOI：
10.1128/mbio.02273-18
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发表时间：
2019-01-01
期刊：
影响因子：
6.4
通讯作者：
Crosson, Sean
中科院分区：
文献类型：
--
作者：
Hershey, David M.;Fiebig, Aretha;Crosson, Sean
由于它们与环境的密切物理作用，表面多糖是细菌适合性的关键决定因素。新月形杆菌在它的一个细胞极上产生一种特殊的结构，称为Holdfast，能够附着在表面上。以前的研究表明，Holdfast是由碳水化合物为基础的物质组成的，并确定了Holdfast发育所需的一些基因。然而，关于其化学结构、生物合成基因和调控原理的不完全信息限制了对Holdfast合成机制的理解。我们利用Holdfast的粘附性，在多天的时间过程中对影响棉布附着的基因进行了饱和筛选。利用转座子文库中突变体的时间分布的相似性，我们定义了与干布定殖相关的离散基因簇。固定合成、鞭毛运动、IV型菌毛组装和平稳的脂多糖(SLP)生产是黏附决定因素的关键类别。对这些簇的详细研究使我们能够预测和实验定义Holdfast和SLPS途径中多个未表征基因的功能。此外，我们还发现菌毛和鞭毛分别通过调节Holdfast抑制剂hfiA来控制Holdfast的合成。这份报告定义了一组有助于粘连的基因，其中包括新发现的Holdfast生物合成和附着所需的基因。我们的数据提供了证据，证明Holdfast含有一个复杂的多糖，在重复单元中至少有四个单糖，并强调了细胞极性在调节新月浑藻附着到表面上的中心作用。重要细菌经常在周围环境中遇到生物和非生物材料，他们经常招募特定的行为程序来定植这些材料。附着力是在表面定植的早期步骤。新月形杆菌产生一种叫做持持器的结构，这种结构允许这种微生物附着在表面并在表面定居。为了了解Holdfast是如何产生的，我们进行了全基因组搜索，通过选择不能附着在棉布上的突变体来寻找导致黏附的基因。我们发现了介导表面接触的基因和有助于保持快速发育的基因之间复杂的相互作用。我们的基因选择确定了可能代表产生Holdfast所需的一组全面的基因，为详细描述制造这种特殊粘附素的酶奠定了基础。
Due to their intimate physical interactions with the environment, surface polysaccharides are critical determinants of fitness for bacteria. Caulobacter crescentus produces a specialized structure at one of its cell poles called the holdfast that enables attachment to surfaces. Previous studies have shown that the holdfast is composed of carbohydrate-based material and identified a number of genes required for holdfast development. However, incomplete information about its chemical structure, biosynthetic genes, and regulatory principles has limited progress in understanding the mechanism of holdfast synthesis. We leveraged the adhesive properties of the holdfast to perform a saturating screen for genes affecting attachment to cheesecloth over a multiday time course. Using similarities in the temporal profiles of mutants in a transposon library, we defined discrete clusters of genes with related effects on cheesecloth colonization. Holdfast synthesis, flagellar motility, type IV pilus assembly, and smooth lipopolysaccharide (SLPS) production represented key classes of adhesion determinants. Examining these clusters in detail allowed us to predict and experimentally define the functions of multiple uncharacterized genes in both the holdfast and SLPS pathways. In addition, we showed that the pilus and the flagellum control holdfast synthesis separately by modulating the holdfast inhibitor hfiA. This report defines a set of genes contributing to adhesion that includes newly discovered genes required for holdfast biosynthesis and attachment. Our data provide evidence that the holdfast contains a complex polysaccharide with at least four monosaccharides in the repeating unit and underscore the central role of cell polarity in mediating attachment of C. crescentus to surfaces.IMPORTANCE Bacteria routinely encounter biotic and abiotic materials in their surrounding environments, and they often enlist specific behavioral programs to colonize these materials. Adhesion is an early step in colonizing a surface. Caulobacter crescentus produces a structure called the holdfast which allows this organism to attach to and colonize surfaces. To understand how the holdfast is produced, we performed a genome-wide search for genes that contribute to adhesion by selecting for mutants that could not attach to cheesecloth. We discovered complex interactions between genes that mediate surface contact and genes that contribute to holdfast development. Our genetic selection identified what likely represents a comprehensive set of genes required to generate a holdfast, laying the groundwork for a detailed characterization of the enzymes that build this specialized adhesin.