Direct target network of the Neurospora crassa plant cell wall deconstruction regulators CLR-1, CLR-2, and XLR-1.
Direct target network of the Neurospora crassa plant cell wall deconstruction regulators CLR-1, CLR-2, and XLR-1.
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DOI：
10.1128/mbio.01452-15
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发表时间：
2015-10-13
期刊：
影响因子：
6.4
通讯作者：
Glass NL
中科院分区：
文献类型：
--
作者：
Craig JP;Coradetti ST;Starr TL;Glass NL
真菌对植物细胞的解构需要大量细胞内和细胞外酶的复杂协调。在粗糙脉孢菌（Neurospora crassa）中，已经鉴定了XLR-1、XLR-2和XLR-1是调节植物细胞壁响应可溶性糖降解的关键转录因子。使用组成型活性突变等位基因定义XLR-1调节子，导致半纤维素酶基因在非诱导条件下表达和分泌。为了确定直接受XLR-1、XLR-2和XLR-1调控的基因，我们对这三种转录因子的表位标记构建体进行了染色质免疫沉淀和下一代测序（ChIPseq）。当N. crassa暴露于植物细胞壁材料时，XLR-1、XLR-2和XLR-1分别与其各自调节子中最强诱导基因的启动子结合。这些包括编码CLR-1和CLR-2纤维素酶（CLR-1/CLR-2）的基因的启动子和编码XLR-1半纤维素酶的基因的启动子。在非诱导性条件下，β-1与其调节子结合;然而，这种结合本身并不转化为基因表达和酶分泌。结合基因的基序分析揭示了保守的DNA结合基序，与其在酿酒酵母中的最接近的paraffin，Gal 4p的匹配的fl-2基序。免疫共沉淀研究表明，α-l和α-l-2以同源复合物形式起作用，但不以α-l/α-l-2异源复合物形式起作用。了解真菌调控复杂的植物细胞壁解构途径，响应多种环境信号，通过相互连接的转录电路提供了深入了解真菌/植物相互作用和真核生物的营养传感。这些调节网络的协调优化可能是最佳微生物酶生产所需的。
Fungal deconstruction of the plant cell requires a complex orchestration of a wide array of intracellular and extracellular enzymes. In Neurospora crassa, CLR-1, CLR-2, and XLR-1 have been identified as key transcription factors regulating plant cell wall degradation in response to soluble sugars. The XLR-1 regulon was defined using a constitutively active mutant allele, resulting in hemicellulase gene expression and secretion under noninducing conditions. To define genes directly regulated by CLR-1, CLR-2, and XLR-1, we performed chromatin immunoprecipitation and next-generation sequencing (ChIPseq) on epitope-tagged constructs of these three transcription factors. When N. crassa is exposed to plant cell wall material, CLR-1, CLR-2, and XLR-1 individually bind to the promoters of the most strongly induced genes in their respective regulons. These include promoters of genes encoding cellulases for CLR-1 and CLR-2 (CLR-1/CLR-2) and promoters of genes encoding hemicellulases for XLR-1. CLR-1 bound to its regulon under noninducing conditions; however, this binding alone did not translate into gene expression and enzyme secretion. Motif analysis of the bound genes revealed conserved DNA binding motifs, with the CLR-2 motif matching that of its closest paralog in Saccharomyces cerevisiae, Gal4p. Coimmunoprecipitation studies showed that CLR-1 and CLR-2 act in a homocomplex but not as a CLR-1/CLR-2 heterocomplex. Understanding fungal regulation of complex plant cell wall deconstruction pathways in response to multiple environmental signals via interconnected transcriptional circuits provides insight into fungus/plant interactions and eukaryotic nutrient sensing. Coordinated optimization of these regulatory networks is likely required for optimal microbial enzyme production.