球毛壳菌(Chaetomium globosum)NK102纤维素酶基因及其表达条件

Genes for cellulose-degradation and their expression conditions in Chaetomium globosum NK102

【目的】生物质的利用是当前生物技术研究的一个热点。本小组分离到一株高效降解纤维素球毛壳菌(Chaetomium globosum)NK102,本文拟探索研究此菌的纤维素酶表达系统并寻找影响酶基因表达的关键因素。【方法】通过对NK102测序,本文界定了球毛壳菌NK102的主要纤维素酶编码基因,使用数字基因表达谱升级版(RNA-Seq)的方法得到纤维素酶基因的表达差异,然后观察了营养、物理条件下纤维素酶基因表达和酶活性变化的情况。【结果】发现随着培养时间的延长,纤维素酶基因整体上表达量升高。在所选基因中,外切葡聚糖酶、纤维二糖脱氢酶和内切葡聚糖酶基因(cbh1,cdh和egl1)的表达量最高。糖代谢的负调控因子ACE I和CreA的随时间表达量均降低,而Hap2/3/5复合体的表达量反而升高。之后检测了不同碳源培养基对纤维素酶基因表达量和酶活性的影响,发现葡萄糖为强阻遏因子,纤维二糖为其诱导物,而山梨醇没有影响。特别是,我们发现光照也影响纤维素酶基因的表达,黑暗条件明显抑制酶基因的表达。【结论】转录组学的方法可以初步探索纤维素酶表达的规律,酶基因的表达受到营养、物理条件的影响。本研究为揭示球毛壳菌降解纤维素分子机理和阐释生物质糖代谢途径提供了有用参考。

[Objective]Use of renewable plant biomass is an active area in current biotechnology research. This report explores the cellulose-degradation system and the factors affecting cellulase gene expression in Chaetomium globosum NK102.[Methods]In the sequenced genome,we identified 10 cellulase genes by sequence homology alignment. We used a high-throughput sequencing technology （RNA-Seq） to monitor the differential expression of the genes under different culture conditions.[Results]We observed that cellulase activity increased with time in the fungal cultures.Accordingly,transcription of the genes encoding cellobiohydrolase,cellobiose dehydrogenase and endoglucanase （cbh1, cdh and egl1） was higher than the others. Expression of the transcriptional repressors,ACE I and CreA,decreased with the time,whereas expression of Hap2/3/5 complex was upregulated. In different carbon sources,cellulase activity and their gene transcription were repressed by glucose and were activated by cellobiose. Sorbitol had no significant effect.Interestingly,light affected positively the expression of these cellulase genes. [Conclusion] Differential RNA-seq analysis can make preliminary exploration on the expression regulation of cellulase genes. Expression of the genes in C.globosum was determined by different culture conditions. This study has shown a molecular system of cellulose-degradation in C.globosum and provides information for interpretation of carbohydrate metabolism.