Enhancing the production of cephalosporin C through modulating the autophagic process of Acremonium chrysogenum.
Enhancing the production of cephalosporin C through modulating the autophagic process of Acremonium chrysogenum.
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通过调节Acremonia chrysogenum的自噬过程提高头孢菌素C的产量
DOI：
10.1186/s12934-018-1021-9
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发表时间：
2018-11-13
影响因子：
6.4
通讯作者：
Liu G
中科院分区：
文献类型：
--
作者：
Li H;Hu P;Wang Y;Pan Y;Liu G
背景自噬用于细胞成分的降解和营养物的回收。 Atg8是自噬中的核心蛋白之一，用作自噬检测的标记物。然而，与酿酒酵母相比，人们对丝状真菌的自噬了解甚少。我们之前的研究表明，自噬相关基因 Acatg1 的破坏通过减少 Acremonium chrysogenum 中头孢菌素生物合成蛋白的降解，显着提高了头孢菌素 C 产量，这表明调节自噬过程是增加 A. chrysogenum 抗生素产量的一种有前景的方法。 chrysogenum。 Acatg8可以补充酿酒酵母中的ATG8突变，表明Acatg8是ATG8的功能同源物。显微镜观察表明，荧光标记的AcAtg8定位于A. chrysogenum的细胞质和自噬体中，并且Acatg8的表达是由营养饥饿诱导的。基因破坏和基因互补表明 Acatg8 对于自噬体形成至关重要。 Acatg8 的破坏显着减少了真菌分生孢子的形成并延迟了分生孢子的萌发。 GFP-AcAtg8 的定位表明自噬参与分生孢子萌发的早期阶段。与 Acatg1 类似，Acatg8 的破坏显着提高了头孢菌素 C 的产量。头孢菌素C生物合成酶（异青霉素N合酶PcbC和异青霉素N差向异构酶CefD2）和过氧化物酶体在Acatg8破坏突变体（ΔAcatg8）中积累，这可能是头孢菌素C产量增强的主要原因。然而，ΔAcatg8的生物量在发酵后期急剧下降，表明自噬对于营养匮乏条件下A. chrysogenum细胞的生存至关重要。 Acatg8 的破坏还会导致线粒体积累，这可能会产生更多的活性氧 (ROS)，从而促进真菌死亡。但过早死亡不利于头孢菌素C的生产。为了解决这个问题，将包含受木糖/木聚糖诱导型启动子控制的 Acatg8 的质粒引入到 ΔAcatg8 中。在补充木糖的培养基中，重组菌株的分生孢子和生长恢复到野生型水平，而头孢菌素C的产量即使延长发酵时间也保持在高水平。结论我们的结果表明Acatg8的诱导表达和Acatg8的破坏显着增加了头孢菌素C的产量。这项研究为提高 A. chrysogenum 中头孢菌素 C 的产量提供了一种有前景的方法。
BackgroundAutophagy is used for degradation of cellular components and nutrient recycling. Atg8 is one of the core proteins in autophagy and used as a marker for autophagic detection. However, the autophagy of filamentous fungi is poorly understood compared with that of Saccharomyces cerevisiae. Our previous study revealed that disruption of the autophagy related gene Acatg1 significantly enhanced cephalosporin C yield through reducing degradation of cephalosporin biosynthetic proteins in Acremonium chrysogenum, suggesting that modulation of autophagic process is one promising way to increase antibiotic production in A. chrysogenum.ResultsIn this study, a S. cerevisiae ATG8 homologue gene Acatg8 was identified from A. chrysogenum. Acatg8 could complement the ATG8 mutation in S. cerevisiae, indicating that Acatg8 is a functional homologue of ATG8. Microscope observation demonstrated the fluorescently labeled AcAtg8 was localized in the cytoplasm and autophagosome of A. chrysogenum, and the expression of Acatg8 was induced by nutrient starvation. Gene disruption and genetic complementation revealed that Acatg8 is essential for autophagosome formation. Disruption of Acatg8 significantly reduced fungal conidiation and delayed conidial germination. Localization of GFP-AcAtg8 implied that autophagy is involved in the early phase of conidial germination. Similar to Acatg1, disruption of Acatg8 remarkably enhanced cephalosporin C yield. The cephalosporin C biosynthetic enzymes (isopenicillin N synthase PcbC and isopenicillin N epimerase CefD2) and peroxisomes were accumulated in the Acatg8 disruption mutant (∆Acatg8), which might be the main reasons for the enhancement of cephalosporin C production. However, the biomass of ΔAcatg8 decreased drastically at the late stage of fermentation, suggesting that autophagy is critical for A. chrysogenum cell survival under nutrition deprived condition. Disruption of Acatg8 also resulted in accumulation of mitochondria, which might produce more reactive oxygen species (ROS) which promotes fungal death. However, the premature death is unfavorable for cephalosporin C production. To solve this problem, a plasmid containing Acatg8 under control of the xylose/xylan-inducible promoter was introduced into ∆Acatg8. Conidiation and growth of the recombinant strain restored to the wild-type level in the medium supplemented with xylose, while the cephalosporin C production maintained at a high level even prolonged fermentation.ConclusionsOur results demonstrated inducible expression of Acatg8 and disruption of Acatg8 remarkably increased cephalosporin C production. This study provides a promising approach for yield improvement of cephalosporin C in A. chrysogenum.
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DOI：
10.1158/1940-6207.capr-10-0387
发表时间：
2011-07
期刊：
Cancer prevention research (Philadelphia, Pa.)
影响因子：
--
作者：
Chen HY;White E
通讯作者：
White E
影响因子：
3.7
作者：
Hirata E;Ohya Y;Suzuki K
通讯作者：
Suzuki K
AcstuA 编码 APSES 转录调节因子，参与产黄顶孢霉的分生孢子形成、头孢菌素生物合成和细胞壁完整性。
影响因子：
3
作者：
Hu, Pengjie;Wang, Ying;Liu, Gang
通讯作者：
Liu, Gang
天蓝色链霉菌的代谢工程可提高灵菌红素 (RED) 的产量。
影响因子：
9.1
作者：
Liu, Panpan;Zhu, Hong;Lu, Yinhua
通讯作者：
Lu, Yinhua
影响因子：
3
作者：
Long Nam Nguyen;Bormann, Joerg;Schaefer, Wilhelm
通讯作者：
Schaefer, Wilhelm