de Castro Patrícia A, Chen Chenxi, de Almeida Ricardo Sérgio Couto, Freitas Fernanda Z, Bertolini Maria C, Morais Enyara Rezende, Brown Neil Andrew, Ramalho Leandra Naira Zambelli, Hagiwara Daisuke, Mitchell Thomas K, Goldman Gustavo H
Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, Brazil.
Mol Microbiol. 2014 Nov;94(3):655-74. doi: 10.1111/mmi.12785. Epub 2014 Sep 30.
Aspergillus fumigatus is an opportunistic pathogen and allergen of mammals. Calcium signalling is essential for A. fumigatus pathogenicity and is regulated by the CrzA transcription factor. We used ChIP-seq (Chromatin Immunoprecipitation DNA sequencing) to explore CrzA gene targets in A. fumigatus. In total, 165 potential binding peaks including 102 directly regulated genes were identified, resulting in the prediction of the A[GT][CG]CA[AC][AG] CrzA-binding motif. The 102 CrzA putatively regulated genes exhibited a diverse array of functions. The phkB (Afu3g12530) histidine kinase and the sskB (Afu1g10940) MAP kinase kinase kinase of the HOG (high-osmolarity glycerol response) pathway were regulated by CrzA. Several members of the two-component system (TCS) and the HOG pathway were more sensitive to calcium. CrzA::GFP was translocated to the nucleus upon osmotic stress. CrzA is important for the phosphorylation of the SakA MAPK in response to osmotic shock. The ΔsskB was more sensitive to CaCl2 , NaCl, and paraquat stress, while being avirulent in a murine model of invasive pulmonary aspergillosis. The presence of CaCl2 and osmotic stresses resulted in synergistic inhibition of ΔcrzA and ΔsskB growth. These results suggest there is a genetic interaction between the A. fumigatus calcineurin-CrzA and HOG pathway that is essential for full virulence.
烟曲霉是哺乳动物的一种机会性病原菌和过敏原。钙信号传导对于烟曲霉的致病性至关重要，并受CrzA转录因子调控。我们使用染色质免疫沉淀DNA测序（ChIP-seq）来探究烟曲霉中CrzA基因的靶点。总共鉴定出165个潜在的结合峰，包括102个直接调控的基因，从而预测出A[GT][CG]CA[AC][AG] CrzA结合基序。这102个推测受CrzA调控的基因表现出多种功能。HOG（高渗甘油应答）途径的phkB（Afu3g12530）组氨酸激酶和sskB（Afu1g10940）丝裂原活化蛋白激酶激酶激酶受CrzA调控。双组分系统（TCS）和HOG途径的几个成员对钙更敏感。在渗透胁迫下，CrzA::GFP易位至细胞核。CrzA对于应对渗透压休克时SakA丝裂原活化蛋白激酶的磷酸化很重要。ΔsskB对氯化钙、氯化钠和百草枯胁迫更敏感，而在侵袭性肺曲霉病的小鼠模型中无毒力。氯化钙和渗透胁迫的存在导致对ΔcrzA和ΔsskB生长的协同抑制。这些结果表明，烟曲霉钙调神经磷酸酶-CrzA和HOG途径之间存在遗传相互作用，这对于完全毒力至关重要。
