Janganan Thamarai K, Chen Gongyou, Chen Daliang, Menino João F, Rodrigues Fernando, Borges-Walmsley Maria I, Walmsley Adrian R
School of Biological and Biomedical Sciences, Durham University, Durham, United Kingdom.
Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal; ICVS/3B's - PT Government Associate Laboratory, University of Minho, Braga/Guimarães, Portugal.
PLoS One. 2015 Sep 3;10(9):e0136866. doi: 10.1371/journal.pone.0136866. eCollection 2015.
The human pathogenic fungus Paracoccidioides brasiliensis (Pb) undergoes a morphological transition from a saprobic mycelium to pathogenic yeast that is controlled by the cAMP-signaling pathway. There is a change in the expression of the Gβ-protein PbGpb1, which interacts with adenylate cyclase, during this morphological transition. We exploited the fact that the cAMP-signaling pathway of Saccharomyces cerevisiae does not include a Gβ-protein to probe the functional role of PbGpb1. We present data that indicates that PbGpb1 and the transcriptional regulator PbTupA both bind to the PKA protein PbTpk2. PbTPK2 was able to complement a TPK2Δ strain of S. cerevisiae, XPY5a/α, which was defective in pseudohyphal growth. Whilst PbGPB1 had no effect on the parent S. cerevisiae strain, MLY61a/α, it repressed the filamentous growth of XPY5a/α transformed with PbTPK2, behaviour that correlated with a reduced expression of the floculin FLO11. In vitro, PbGpb1 reduced the kinase activity of PbTpk2, suggesting that inhibition of PbTpk2 by PbGpb1 reduces the level of expression of Flo11, antagonizing the filamentous growth of the cells. In contrast, expressing the co-regulator PbTUPA in XPY5a/α cells transformed with PbTPK2, but not untransformed cells, induced hyperfilamentous growth, which could be antagonized by co-transforming the cells with PbGPB1. PbTUPA was unable to induce the hyperfilamentous growth of a FLO8Δ strain, suggesting that PbTupA functions in conjunction with the transcription factor Flo8 to control Flo11 expression. Our data indicates that P. brasiliensis PbGpb1 and PbTupA, both of which have WD/β-propeller structures, bind to PbTpk2 to act as antagonistic molecular switches of cell morphology, with PbTupA and PbGpb1 inducing and repressing filamentous growth, respectively. Our findings define a potential mechanism for controlling the morphological switch that underpins the virulence of dimorphic fungi.
人类致病真菌巴西副球孢子菌（Pb）会经历从腐生菌丝体到致病酵母的形态转变，这一过程受cAMP信号通路控制。在此形态转变过程中，与腺苷酸环化酶相互作用的Gβ蛋白PbGpb1的表达发生了变化。我们利用酿酒酵母的cAMP信号通路不包含Gβ蛋白这一事实，来探究PbGpb1的功能作用。我们提供的数据表明，PbGpb1和转录调节因子PbTupA都与PKA蛋白PbTpk2结合。PbTPK2能够补充酿酒酵母TPK2Δ菌株XPY5a/α中假菌丝生长缺陷。虽然PbGPB1对亲本酿酒酵母菌株MLY61a/α没有影响，但它抑制了用PbTPK2转化的XPY5a/α的丝状生长，这种行为与絮凝蛋白FLO11表达的降低相关。在体外，PbGpb1降低了PbTpk2的激酶活性，表明PbGpb1对PbTpk2的抑制降低了Flo11的表达水平，拮抗了细胞的丝状生长。相反，在已用PbTPK2转化但未转化的XPY5a/α细胞中表达共调节因子PbTUPA，会诱导过度丝状生长，而用PbGPB1共转化细胞可拮抗这种生长。PbTUPA无法诱导FLO8Δ菌株的过度丝状生长，这表明PbTupA与转录因子Flo8协同作用来控制Flo11的表达。我们的数据表明，巴西副球孢子菌的PbGpb1和PbTupA都具有WD/β-螺旋桨结构，它们与PbTpk2结合，作为细胞形态的拮抗分子开关，PbTupA和PbGpb1分别诱导和抑制丝状生长。我们的研究结果确定了一种控制形态转变的潜在机制，这种形态转变是双态真菌毒力的基础。
