木聚糖协同纤维素诱导纤维素酶的激发效应及其机制

The cellulolytic enzymes play an important role in soil carbon cycling and agricultural waste utilization (composting), so how to improve the efficiency of this bioprocess regulated by fungal cellulases has obtained greatly attentions in the world. Our preliminary results found that the cellulase activity was low when the straw-degrading fungus Penicillium oxalicum GZ-2 induced by cellulose or xylan only. The cellulase activity increases nearly 8 times when P. oxalicum GZ-2 was induced by the mixture of cellulose and xylan, which was defined as "priming effect" of cellulase production in this study. However, the synergistic mechanisms of priming effect are still unknown. In this project, the influence of concentration of mixed substances on priming effect will be evaluated, and the signaling molecules activated the priming effect will be analyzed using metabolomics. Synergistic regulation mechanisms of priming effect induced by the mixed substrates in P. oxalicum will be analyzed using RNA-seq and iTRAQ. The relationship between of transcriptional regulator genes (creA and xlnR) and priming effect will be evaluated, and their genes disruption by homologous recombination will be performed with the plasmid pDHt/xlnR::hph. Comparative analysis of the extracellular proteins and digital gene expression profile inoculated by wide type strain and the mutants will be carried out under different induced condition. These studies will provide theoretical instructions and technical supports for improving the cellulolytic enzymes production induced by complex substrates, and also provide a theoretical basis to improve the utilization efficiency of agricultural wastes.

纤维素酶在土壤碳循环和堆肥腐熟过程中发挥着重要作用，调控纤维素酶高效表达已成为国内外研究热点。前期研究发现当纤维素或木聚糖单独诱导秸秆降解菌草酸青霉GZ-2时纤维素酶活力均较低，而两者混合诱导时酶活力提高近8倍，即产生木聚糖协同纤维素诱导纤维素酶的激发效应。然而，两者诱导调控激发效应的机制尚不清楚。本项目以菌株GZ-2为材料，研究木聚糖与纤维素相对浓度对激发效应的影响，明确两者最佳诱导比例与相对贡献；应用转录组学与稳定同位素标记蛋白组学（iTRAQ）解析激发效应下纤维素酶高效表达机制；分析转录调控因子（creA 和 xlnR）与激发效应的关系，构建二元载体定点敲除creA和xlnR基因，比较分析野生菌株和突变菌株对激发效应的响应差异，揭示它们在激发效应中的调控机制，为农业废弃物资源化高效利用提供科学依据。

微生物分泌的纤维素酶在土壤碳循环和堆肥腐熟过程中发挥着重要作用，木质纤维素类底物调控纤维素酶高效表达已成为国内外研究热点。本项目以前期分离的水稻秸秆高效降解菌株草酸青霉GZ-2为材料，前期研究发现秸秆类底物可以高效诱导GZ-2产木质纤维素酶，而以纯纤维素或木聚糖单独诱导时GZ-2时分泌纤维素酶活力均较低，而两者混合诱导时酶活力提高近8倍，即产生木聚糖协同纤维素诱导纤维素酶的激发效应。然而，木聚糖与纤维素协调诱导调控激发效应的机制尚不清楚。本项目以草酸青霉GZ-2为材料，通过应用分子生物学，蛋白质组学等技术系统地研究木聚糖与纤维素相对浓度对激发效应的影响，明确两者最佳诱导比例与相对贡献；应用蛋白组学解析激发效应下纤维素酶高效表达机制；克隆表达了多个木质纤维素酶基因，解析了这些基因表达模式与激发效应的关系，揭示它们在激发效应中的作用机制，为农业废弃物资源化高效利用提供科学依据。