Targeted changes of the cell wall proteome influence Candida albicans ability to form single- and multi-strain biofilms.
Targeted changes of the cell wall proteome influence Candida albicans ability to form single- and multi-strain biofilms.
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细胞壁蛋白质组的靶向变化会影响白色念珠菌形成单晶和多晶粒生物膜的能力。
DOI：
10.1371/journal.ppat.1004542
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发表时间：
2014-12
期刊：
影响因子：
6.7
通讯作者：
d'Enfert C
中科院分区：
文献类型：
--
作者：
Cabral V;Znaidi S;Walker LA;Martin-Yken H;Dague E;Legrand M;Lee K;Chauvel M;Firon A;Rossignol T;Richard ML;Munro CA;Bachellier-Bassi S;d'Enfert C
生物膜形成是致病性酵母菌白念珠菌的重要毒力特征。我们结合基因过表达、菌株条形码和微阵列分析来筛选531 C文库。白念珠菌条件性过表达菌株（基因组的约10%）中影响混合群体实验中生物膜发育的基因。16个基因的过表达增加了菌株在多菌株生物膜中的占有率，而4个基因的过表达降低了它，这16个基因的集合显著富集了那些编码预测的糖基磷脂酰肌醇（GPI）修饰蛋白的基因，即Ihd 1/Pga 36、Phr 2、Pga 15、Pga 19、Pga 22、Pga 32、Pga 37、Pga 42和Pga 59;其中八种被归类为病原体特异性。使用单独或竞争性生长的过表达菌株的验证实验表明，这些基因对生物膜形成的贡献是可变的和阶段特异性的。使用原子力显微镜在单细胞分辨率下对PGA 59和PGA 22进行更深入的功能分析，结果表明，任何一个基因的过表达都增加了C。白色念珠菌粘附于非生物基质的能力。然而，与PGA 59不同，PGA 22过表达导致细胞簇形成，导致对剪切力的敏感性增加和形成单菌株生物膜的能力降低。在由PGA 22非过表达细胞提供的多菌株环境中，保护PGA 22过表达细胞免受剪切力并且更适合生物膜发育。超微结构分析，全基因组转录谱和表型分析在异源背景下表明，PGA 22通过改变细胞壁结构和/或功能影响细胞粘附。综上所述，我们的研究结果表明，一些新的预测GPI修饰的蛋白质有助于生物膜细胞之间的合作行为，是C。白色念珠菌生物膜形成。此外，他们说明了使用签名标签结合基因过表达的能力，用于鉴定涉及与C有关的过程的新基因。白色念珠菌毒力 白色念珠菌是最常见的人类真菌病原体。它引起疾病的能力部分依赖于生物膜的形成，这是一种对抗真菌剂耐受的高度粘附细胞的保护结构和宿主免疫反应。生物膜被认为是感染的持久根源，将感染细胞传播到其他位置。在这项研究中，我们进行了大规模的表型分析，旨在确定基因的过度表达影响生物膜的发展，在C。白色念珠菌。我们的屏幕依赖于531 C的集合。白色念珠菌菌株，每个菌株条件性过表达一个给定的基因，并携带一个特定的分子标签，允许在混合群体实验中定量菌株丰度。我们的研究结果惊人地揭示了在531株含生物膜模型中，过度产生被称为Pgas（推定GPI锚定蛋白）的特征较差的表面蛋白的菌株的富集。我们发现，这些PGA基因差异有助于单菌株和多菌株生物膜的形成，并参与生物膜发育过程的特定阶段。综上所述，我们的结果揭示了C。白念珠菌细胞表面蛋白在生物膜形成过程中的变化，并反映了菌株条形码与基因过表达相结合的强大用途，以鉴定与病原微生物毒力有关的过程中涉及的基因和/或途径。
Biofilm formation is an important virulence trait of the pathogenic yeast Candida albicans. We have combined gene overexpression, strain barcoding and microarray profiling to screen a library of 531 C. albicans conditional overexpression strains (∼10% of the genome) for genes affecting biofilm development in mixed-population experiments. The overexpression of 16 genes increased strain occupancy within a multi-strain biofilm, whereas overexpression of 4 genes decreased it. The set of 16 genes was significantly enriched for those encoding predicted glycosylphosphatidylinositol (GPI)-modified proteins, namely Ihd1/Pga36, Phr2, Pga15, Pga19, Pga22, Pga32, Pga37, Pga42 and Pga59; eight of which have been classified as pathogen-specific. Validation experiments using either individually- or competitively-grown overexpression strains revealed that the contribution of these genes to biofilm formation was variable and stage-specific. Deeper functional analysis of PGA59 and PGA22 at a single-cell resolution using atomic force microscopy showed that overexpression of either gene increased C. albicans ability to adhere to an abiotic substrate. However, unlike PGA59, PGA22 overexpression led to cell cluster formation that resulted in increased sensitivity to shear forces and decreased ability to form a single-strain biofilm. Within the multi-strain environment provided by the PGA22-non overexpressing cells, PGA22-overexpressing cells were protected from shear forces and fitter for biofilm development. Ultrastructural analysis, genome-wide transcript profiling and phenotypic analyses in a heterologous context suggested that PGA22 affects cell adherence through alteration of cell wall structure and/or function. Taken together, our findings reveal that several novel predicted GPI-modified proteins contribute to the cooperative behaviour between biofilm cells and are important participants during C. albicans biofilm formation. Moreover, they illustrate the power of using signature tagging in conjunction with gene overexpression for the identification of novel genes involved in processes pertaining to C. albicans virulence. Candida albicans is the most prevalent human fungal pathogen. Its ability to cause disease relies, in part, on the formation of biofilms, a protective structure of highly adherent cells tolerant to antifungal agents and the host immune response. The biofilm is considered as a persistent root of infection, disseminating infectious cells to other locations. In this study, we performed large-scale phenotypic analyses aimed at identifying genes whose overexpression affects biofilm development in C. albicans. Our screen relied on a collection of 531 C. albicans strains, each conditionally overexpressing one given gene and carrying one specific molecular tag allowing the quantification of strain abundance in mixed-population experiments. Our results strikingly revealed the enrichment of strains overproducing poorly-characterized surface proteins called Pgas (Putative GPI-Anchored proteins), within a 531-strain-containing biofilm model. We show that these PGA genes differentially contribute to single-strain and multi-strain biofilm formation and are involved in specific stages of the biofilm developmental process. Taken together, our results reveal the importance of C. albicans cell surface proteins during biofilm formation and reflect the powerful use of strain barcoding in combination with gene overexpression to identify genes and/or pathways involved in processes pertaining to virulence of pathogenic microbes.
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