Disruption of gul-1 decreased the culture viscosity and improved protein secretion in the filamentous fungus Neurospora crassa.
Disruption of gul-1 decreased the culture viscosity and improved protein secretion in the filamentous fungus Neurospora crassa.
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gul-1 的破坏降低了丝状真菌粗糙脉孢菌中的培养物粘度并改善了蛋白质分泌
DOI：
10.1186/s12934-018-0944-5
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发表时间：
2018-06-16
影响因子：
6.4
通讯作者：
Tian C
中科院分区：
文献类型：
--
作者：
Lin L;Sun Z;Li J;Chen Y;Liu Q;Sun W;Tian C
背景：纤维素分解真菌粗糙脉孢菌被认为是生产酶和生物乙醇的潜在宿主。然而，大规模应用受到其丝状生长的阻碍。虽然以前的调查表明，菌丝体形态在液体培养可以通过改变物理因素进行控制，有很少的知识可用于形态控制的遗传modification.Results的潜力：在这项研究中，我们筛选的丝状真菌N.粗鲁。在筛选的90个形态突变体中，14个突变体表现出比野生型菌株高得多的粘度，只有两个突变体在深层培养中表现出低粘度的形态。我们观察到，gul-1（NCU 01197），它编码的mRNA结合蛋白参与细胞壁重塑，破坏引起沉淀形成的发酵进展，并导致最显着的粘度降低培养液。此外，gul-1的过量表达导致粘度显著增加，表明gul-1在液体培养过程中对菌丝形态具有重要作用。转录谱分析显示，编码八种GPI锚定细胞壁蛋白的基因在Δgul-1中的表达降低，而与两种非锚定细胞壁蛋白相关的基因的表达升高。同时，两个疏水蛋白基因的表达水平也发生了显著变化。这些结果表明，GUL-1影响细胞壁相关基因的转录，从而影响细胞壁结构和菌丝形态。此外，gul-1的缺失导致蛋白质分泌增加，可能是由于细胞壁完整性的缺陷，这表明这是用于酶生产的菌株改良的替代策略。为了证实实际应用，在超纤维素酶产生菌（nncw-1）Ncap 3 m中缺失gul-1显著降低了培养液的粘度。crassa的形态突变体，探索影响菌丝形态的基因。破坏几个候选基因改变了液体培养中的粘度。本研究为利用基因工程技术控制深层发酵中真菌的形态提供了一个范例，也为工业真菌菌种的改良提供了有益的参考。
Background:The cellulolytic fungus Neurospora crassa is considered a potential host for enzyme and bioethanol production. However, large scale applications are hindered by its filamentous growth. Although previous investigations have shown that mycelial morphology in submerged culture can be controlled by altering physical factors, there is little knowledge available about the potential for morphology control by genetic modification.Results:In this study, we screened morphological mutants in the filamentous fungus N. crassa. Of the 90 morphological mutants screened, 14 mutants exhibited considerably higher viscosity compared with that of the wild type strain, and only two mutants showed low-viscosity morphologies in submerged culture. We observed that disruption of gul-1 (NCU01197), which encodes an mRNA binding protein involved in cell wall remodeling, caused pellet formation as the fermentation progressed, and resulted in the most significant decrease in viscosity of culture broth. Moreover, over-expression of gul-1 caused dramatically increased viscosity, suggesting that the gul-1 had an important function in mycelial morphology during submerged cultivation. Transcriptional profiling showed that expression of genes encoding eight GPI-anchored cell wall proteins was lowered in Δgul-1 while expression of genes associated with two non-anchored cell wall proteins was elevated. Meanwhile, the expression levels of two hydrophobin genes were also significantly altered. These results suggested that GUL-1 affected the transcription of cell wall-related genes, thereby influencing cell wall structure and mycelial morphology. Additionally, the deletion of gul-1 caused increased protein secretion, probably due to a defect in cell wall integrity, suggesting this as an alternative strategy of strain improvement for enzyme production. To confirm practical applications, deletion of gul-1 in the hyper-cellulase producing strain (∆ncw-1∆Ncap3m) significantly reduced the viscosity of culture broth.Conclusions:Using the model filamentous fungus N. crassa, genes that affect mycelial morphology in submerged culture were explored through systematic screening of morphological mutants. Disrupting several candidate genes altered viscosities in submerged culture. This work provides an example for controlling fungal morphology in submerged fermentation by genetic engineering, and will be beneficial for industrial fungal strain improvement.
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DOI：
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发表时间：
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期刊：
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