Analysis of Candida albicans mutants defective in the Cdk8 module of mediator reveal links between metabolism and biofilm formation.
Analysis of Candida albicans mutants defective in the Cdk8 module of mediator reveal links between metabolism and biofilm formation.
复制标题
DOI：
10.1371/journal.pgen.1004567
复制
发表时间：
2014-10
期刊：
影响因子：
4.5
通讯作者：
Hogan DA
中科院分区：
文献类型：
--
作者：
Lindsay AK;Morales DK;Liu Z;Grahl N;Zhang A;Willger SD;Myers LC;Hogan DA
白色念珠菌生物膜形成是一个关键的毒力特征，涉及菌丝生长和粘附素表达。绿脓素 (PYO) 是一种由铜绿假单胞菌分泌的吩嗪，可抑制白色念珠菌生物膜的形成和皱纹菌落的发育。通过基因筛选，我们鉴定了两个突变体：ssn3Δ/Δ 和 ssn8Δ/Δ，它们在 PYO 存在的情况下继续起皱。 Ssn8是一种细胞周期蛋白样蛋白，Ssn3类似于细胞周期蛋白依赖性激酶；这两种蛋白都是异四聚体 Cdk8 模块的一部分，该模块与转录共调节因子 Mediator 形成复合物。 Ssn3 激酶活性对于 PYO 敏感性也是必需的，因为激酶死亡突变体在 PYO 存在的情况下仍保持皱纹集落形态。此外，在缺乏 Cdk8 模块的其他两个组件（Srb8 和 Srb9）的突变体中观察到类似的表型。通过代谢组学分析和生化测定，我们发现，即使在 PYO 存在的情况下，受损的 Cdk8 模块也会导致葡萄糖消耗、糖酵解相关转录本、氧化代谢和 ATP 水平增加。在突变体中，呼吸抑制达到与 PYO 处理的野生型相当的水平，抑制了皱纹集落的发育。多项证据表明 PYO 并不通过 Cdk8 发挥作用。最后，ssn3突变体是一种超生物膜形成者，并且在PYO存在的情况下保持比野生型更高的生物膜形成。这些数据共同提供了关于 Mediator 的 Cdk8 模块在调节白色念珠菌生理学中的作用以及呼吸活动与皱纹菌落和生物膜发育之间的联系的新见解。 白色念珠菌目前是医院感染最常见的原因之一，可引起从鹅口疮到危及生命的全身感染等疾病。白色念珠菌很容易在植入设备（例如导管和假牙）上形成生物膜，而生物膜与全身感染和抗真菌药物耐药性风险增加有关。我们之前表明生物膜的形成与氧化代谢呈正相关。在这里，我们发现，当转录共调节因子 Mediator 的 Cdk8 模块受到损害时，呼吸、皱纹集落形成、生物膜形成和对绿脓素抑制作用的抵抗力都会增加。 Cdk8模块突变体都表现出基础代谢的差异，这表明代谢活性增加。此外，使用这些菌株，我们发现在不存在生长缺陷的情况下呼吸活动的抑制与生物膜形成的抑制之间存在直接相关性，支持白色念珠菌代谢和形态相关的模型。目前正在研究人类 Cdk8 模块作为癌症化疗的靶点，因此需要了解抑制该模块的后果或潜在益处。
Candida albicans biofilm formation is a key virulence trait that involves hyphal growth and adhesin expression. Pyocyanin (PYO), a phenazine secreted by Pseudomonas aeruginosa, inhibits both C. albicans biofilm formation and development of wrinkled colonies. Using a genetic screen, we identified two mutants, ssn3Δ/Δ and ssn8Δ/Δ, which continued to wrinkle in the presence of PYO. Ssn8 is a cyclin-like protein and Ssn3 is similar to cyclin-dependent kinases; both proteins are part of the heterotetrameric Cdk8 module that forms a complex with the transcriptional co-regulator, Mediator. Ssn3 kinase activity was also required for PYO sensitivity as a kinase dead mutant maintained a wrinkled colony morphology in the presence of PYO. Furthermore, similar phenotypes were observed in mutants lacking the other two components of the Cdk8 module—Srb8 and Srb9. Through metabolomics analyses and biochemical assays, we showed that a compromised Cdk8 module led to increases in glucose consumption, glycolysis-related transcripts, oxidative metabolism and ATP levels even in the presence of PYO. In the mutant, inhibition of respiration to levels comparable to the PYO-treated wild type inhibited wrinkled colony development. Several lines of evidence suggest that PYO does not act through Cdk8. Lastly, the ssn3 mutant was a hyperbiofilm former, and maintained higher biofilm formation in the presence of PYO than the wild type. Together these data provide novel insights into the role of the Cdk8 module of Mediator in regulation of C. albicans physiology and the links between respiratory activity and both wrinkled colony and biofilm development. Candida albicans is currently one of the most common causes of nosocomial infections, and causes diseases ranging from oral thrush to life-threatening systemic infections. C. albicans readily forms biofilms on implanted devices, such as catheters and dentures, and biofilms are associated with increased risk of systemic infections and resistance to antifungals. We previously showed that biofilm formation positively correlates with oxidative metabolism. Here, we show that respiration, wrinkled colony formation, biofilm formation, and resistance to the inhibitory effects of pyocyanin were increased when the Cdk8 module of Mediator, a co-regulator of transcription, was compromised. Cdk8 module mutants all exhibited differences in basal metabolism that were indicative of increased metabolic activity. Furthermore, using these strains, we showed a direct correlation between the inhibition of respiratory activity, in the absence of growth defects, and inhibition of biofilm formation, supporting a model in which C. albicans metabolism and morphology are linked. The human Cdk8 module is currently being investigated as a target for cancer chemotherapy, and thus an understanding of the consequences or potential benefits of inhibiting this module is required.
登录
查看更多内容
影响因子：
6.7
作者：
Blankenship JR;Fanning S;Hamaker JJ;Mitchell AP
通讯作者：
Mitchell AP
影响因子：
3.9
作者：
Fan, JJ;Chaturvedi, V;Shen, SH
通讯作者：
Shen, SH
影响因子：
3
作者：
Baillie, GS;Douglas, LJ
通讯作者：
Douglas, LJ
影响因子：
3.7
作者：
Ayer A;Sanwald J;Pillay BA;Meyer AJ;Perrone GG;Dawes IW
通讯作者：
Dawes IW
影响因子：
4.8
作者：
Borggrefe, T;Davis, R;Kornberg, RD
通讯作者：
Kornberg, RD