Wang Juan, Gong Yanfen, Zhao Shengming, Liu Gang
1Shenzhen Key Laboratory of Microbial Genetic Engineering, College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, People's Republic of China.
Shenzhen Key Laboratory of Marine Bioresources and Ecology, Shenzhen, 518060 People's Republic of China.
3 Biotech. 2018 Mar;8(3):160. doi: 10.1007/s13205-017-1069-y. Epub 2018 Mar 5.
(ATCC 42464) is a thermophilic fungus that produces cellulolytic enzymes with high thermal stability. Unlike its mesophile counterparts, study on gene expression regulation of cellulolytic enzymes in is inadequate. This work identified the function of MHR1, a putative transcription regulator of cellulolytic enzymes in that was found through RNA-Seq based gene expression profile analysis. RNA interference was used to study the role of MHR1. A recombinant plasmid, pUC19-P--T, which contained the RNAi sequence for was constructed and transformed into . One of the transformants, MtR5, in which the RNA interference efficiency was the highest, was used for the following studies. In the -silenced strain MtR5, the filter paper hydrolyzing activity was 1.33-fold; β-1, 4-endoglucanase activity was 1.65-fold; and xylanase activity was 1.48-fold higher than those of the parental strain after induction, respectively, by wheat straw powder. qRT-PCR showed that gene expression of , , and were 9.56-, 37.36-, 56.14- and 28.30-fold higher in MtR5 than in wild type, respectively. Our findings suggest that the transcription factor MHR1 of can repress cellulase and xylanase activities. Silenced results in increased expression not only of the main cellulase genes, but also of the positive regulatory gene . This work is relevant to the development of as an industrial production host for cellulolytic and hemicellulolytic enzymes, which could be used to degrade a wide range of different biomass, converting lignocellulosic feedstock into sugar precursors for biofuels.
（ATCC 42464）是一种嗜热真菌，可产生具有高热稳定性的纤维素分解酶。与其嗜温同类真菌不同，对其纤维素分解酶基因表达调控的研究尚不充分。本研究通过基于RNA测序的基因表达谱分析，鉴定了嗜热栖热放线菌中一种假定的纤维素分解酶转录调节因子MHR1的功能。采用RNA干扰技术研究MHR1的作用。构建了包含嗜热栖热放线菌RNA干扰序列的重组质粒pUC19 - P - - T，并将其转化至嗜热栖热放线菌中。选择RNA干扰效率最高的转化子之一MtR5用于后续研究。在MHR1沉默菌株MtR5中，经麦草粉诱导后，滤纸水解活性比亲本菌株高1.33倍；β - 1,4 - 内切葡聚糖酶活性高1.65倍；木聚糖酶活性高1.48倍。qRT - PCR结果显示，MtR5中cbh1、cbh2、egl1和xyn1的基因表达分别比野生型高9.56、37.36、56.14和28.30倍。我们的研究结果表明，嗜热栖热放线菌的转录因子MHR1可抑制纤维素酶和木聚糖酶的活性。MHR1沉默不仅导致主要纤维素酶基因的表达增加，还导致正调控基因xyr1的表达增加。本研究对于嗜热栖热放线菌作为纤维素分解酶和半纤维素分解酶工业生产宿主的开发具有重要意义，这些酶可用于降解多种不同的生物质，将木质纤维素原料转化为生物燃料的糖前体。
