Transcriptomic profiling and genetic analyses reveal novel key regulators of cellulase and xylanase gene expression in Penicillium oxalicum.
Transcriptomic profiling and genetic analyses reveal novel key regulators of cellulase and xylanase gene expression in Penicillium oxalicum.
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转录组分析和遗传分析揭示了草酸青霉中纤维素酶和木聚糖酶基因表达的新关键调节因子
DOI：
10.1186/s13068-017-0966-y
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发表时间：
2017
影响因子：
6.3
通讯作者：
Feng JX
中科院分区：
文献类型：
--
作者：
Yan YS;Zhao S;Liao LS;He QP;Xiong YR;Wang L;Li CX;Feng JX
背景：向更加环境友好型经济的过渡促进了对现代生物精炼的研究，包括利用低价值的木质纤维素。生物炼制的广泛应用所面临的主要挑战是能够有效地将柠檬酸纤维素水解成糖的酶的高成本。青霉产生大量的植物细胞壁降解酶，但是它们的产生受到复杂的调控网络的严格控制，导致天然酶的产量低。调控基因已成为基因工程的目标，以提高微生物中的酶的生产。在这项研究中，我们使用转录组分析和遗传分析，筛选和确定新的关键调控纤维素酶和木聚糖酶基因表达的P。结果：对不同碳源（包括葡萄糖、麦麸和麦麸加Avicel）上的P.acericumHP7 -1的转录组进行比较分析，确定了40个调节纤维素分解酶基因表达的候选基因。对31个候选基因进行缺失突变，得到11个滤纸纤维素酶产量与出发菌株P.poxKu70相比发生显著变化的突变株。在这11个突变体中，ΔPoxCxrA、ΔPoxCxrB和Δ PoxNsdD显示出最显著的酶产量降低（分别为96.8%、75.9%和58.5%）。本文首次报道了这11个基因中的10个与纤维素酶的产生有关。进一步的试验表明，ΔPoxCxrA、ΔPoxCxrB和Δ PoxNsdD均能显著降低木聚糖酶的产量，而Δ PoxCxrA的木聚糖酶产量可忽略不计。有趣的是，Δ PoxCxr和Δ PoxNsdD显示出显著增加的β-葡萄糖苷酶产量。实时定量逆转录-PCR和电泳迁移率变动分析（EMSA）表明，在纤维素存在下，真菌细胞生长过程中，PoxCxrA、PoxCxrB和PoxNsdD相互调控，但调控模式动态变化。EMSA表明，PoxCxrA，PoxCxrB，和PoxNsdD直接结合推定的主要纤维素酶和木聚糖酶基因的启动子。结论：我们已经检测到并确定了三个关键的新的调控基因，PoxCxrA，PoxCxrB，和PoxNsdD，直接和间接调节纤维素酶和木聚糖酶基因的表达在P. aceticum。本研究为真菌纤维素酶和木聚糖酶基因表达调控机制提供了新的见解。
Background:The transition to a more environmentally friendly economy has prompted studies of modern biorefineries, including the utilization of low-value lignocellulose. The major challenge facing the widespread application of biorefineries is the high cost of enzymes that can efficiently hydrolyze recalcitrant cellulose to sugars.Penicillium oxalicumproduces large amounts of plant-cell-wall-degrading enzymes, but their production is tightly controlled by complex regulatory networks, resulting in low yields of the native enzymes. Regulatory genes have been the targets of genetic engineering to improve enzyme production in microorganisms. In this study, we used transcriptomic profiling and genetic analyses to screen for and identify novel key regulators of cellulase and xylanase gene expression inP. oxalicum.Results:A comparative analysis of the transcriptomes ofP.oxalicumHP7-1 on different carbon sources, including glucose, wheat bran, and wheat bran plus Avicel, identified 40 candidate genes regulating the expression of cellulolytic enzyme genes. Deletion mutants of 31 candidate genes were constructed inP.oxalicum∆PoxKu70and 11 resultant mutants showed significant changes in their filter-paper cellulase production compared with the parental strain ∆PoxKu70. Among these 11 mutants, ΔPoxCxrA, ΔPoxCxrB, and ΔPoxNsdDshowed the most significant reduction in the enzyme production (96.8, 75.9, and 58.5%, respectively). Ten of these 11 genes are here reported to be involved in cellulase production for the first time. Further tests revealed that ΔPoxCxrA, ΔPoxCxrB, and ΔPoxNsdDdisplayed significantly reduced xylanase production, whereas ΔPoxCxrAproduced negligible xylanase. Interestingly, ΔPoxCxrBand ΔPoxNsdDshowed significantly increased β-glucosidase production. Real-time quantitative reverse transcription-PCR and an electrophoretic mobility shift assay (EMSA) showed thatPoxCxrA,PoxCxrB, andPoxNsdDregulate the expression of one another, but the mode of regulation changes dynamically during the growth of fungal cells in the presence of cellulose. EMSA showed that PoxCxrA, PoxCxrB, and PoxNsdD directly bind the putative promoters of major cellulase and xylanase genes.Conclusions:We have detected and identified three key new regulatory genes,PoxCxrA,PoxCxrB, andPoxNsdD, that directly and indirectly regulate the expression of cellulase and xylanase genes inP.oxalicum. This study provides novel insights into the regulatory mechanisms of fungal cellulase and xylanase gene expression.
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影响因子：
48
作者：
Huber W;Carey VJ;Gentleman R;Anders S;Carlson M;Carvalho BS;Bravo HC;Davis S;Gatto L;Girke T;Gottardo R;Hahne F;Hansen KD;Irizarry RA;Lawrence M;Love MI;MacDonald J;Obenchain V;Oleś AK;Pagès H;Reyes A;Shannon P;Smyth GK;Tenenbaum D;Waldron L;Morgan M
通讯作者：
Morgan M
影响因子：
48
作者：
Langmead, Ben;Salzberg, Steven L.
通讯作者：
Salzberg, Steven L.
影响因子：
6.3
作者：
Häkkinen M;Valkonen MJ;Westerholm-Parvinen A;Aro N;Arvas M;Vitikainen M;Penttilä M;Saloheimo M;Pakula TM
通讯作者：
Pakula TM
影响因子：
5
作者：
Noguchi, Yuji;Sano, Motoaki;Kobayashi, Tetsuo
通讯作者：
Kobayashi, Tetsuo
影响因子：
3.6
作者：
Han, KH;Han, KY;Han, DM
通讯作者：
Han, DM