华根霉液态培养生产脂肪酶不同形态菌体的转录组差异分析

Transcriptome comparison of Rhizopus chinensis mycelia with different morphology in submerged fermentation for lipase production

摘要:

【背景】作为发酵工业中一类重要的生产菌株，丝状真菌目的产物的形成与菌体形态有着紧密的联系。华根霉(Rhizopus chinensis) CCTCC M201021是从我国传统酿造浓香型白酒大曲中筛选到的一株丝状真菌，其生成的包括脂肪酶在内的酶蛋白具有较高的工业应用价值。【目的】华根霉(Rhizopus chinensis) CCTCC M201021在脂肪酶液态发酵中形成两种不同形态菌体，发酵表现差异明显。本研究考察华根霉不同菌体形态及其细胞代谢在转录组水平的内在差异。【方法】基于RNA-Seq高通量转录组测序，分别对液态培养获得的不同形态华根霉菌体高表达和显著差异表达的转录基因进行功能分析。【结果】两种形态菌体转录组存在明显的差异。利用RPKM值对表达量最高的前20个基因进行分析，聚集态菌体高表达基因主要为不同类别的核糖体蛋白，而分散态菌体中与细胞形态相关的几丁质酶及与信号传导相关的基因也是高表达基因。在两种形态菌体显著差异表达的20个基因中，除了涉及代谢的基因有明显不同外，分散态菌体中也有一些涉及“细胞过程与信号”的基因上调表达显著。两种形态菌体中独有表达基因总体表达量均较低，但聚集态菌体独有表达基因在基因种类和表达量上都要明显高于分散态菌体。同时，转录分析表明，华根霉脂肪酶在聚集菌体中较高的生产水平与脂肪酶基因的高水平转录有关。【结论】菌体形态的差异显著影响了华根霉的转录组，不仅不同形态菌体高表达基因和显著差异表达基因有明显不同，而且功能相同的蛋白在不同菌体形态下也多是由不同基因表达，它们可能承担着不同的作用。总体而言，华根霉聚集态菌体中存在更为复杂的生理过程，而分散菌体中受到信号的传导和调控似乎更多。菌体形态的改变可能是细胞分化的结果，伴随着菌体对细胞微环境改变的一种响应。研究结果为深入了解丝状真菌形态分化的内在机制及其影响提供了一些线索。

Abstract:

[Background] Filamentous fungi are important production strains in fermentation industry, and their morphology is related with the production of target products. Rhizopus chinensis CCTCC M201021 is an important filamentous fungus isolated from Daqu for Chinese liquor, produces some important industrial enzymes, such as lipases. [Objective] Rhizopus chinensis CCTCC M201021 can form two typical mycelia morphology, the aggregated and dispersed, in submerged culture with different fermentation performance. The objective of this work was to study the intrinsic difference associated with different mycelia morphology based on transcriptome profiles of R. chinensis. [Methods] Based on high throughput RNA sequencing, gene transcriptions of high expression and significantly differential expression from R. chinensis mycelia with different morphology were compared. [Results] RNA sequencing released the significant difference in the transcriptomes of mycelia with different morphology. In the 20 genes featuring the highest RPKM values in the transcriptome of the aggregated or dispersed mycelia, most genes with the highest RPKM value were ribosomal proteins genes in the aggregated mycelia. Chitinase genes and signal transduction genes were found in the dispersed R. chinensis in addition to some ribosomal proteins genes. Analysis of 20 most up-regulated genes in the transcriptome of the aggregated or dispersed mycelia indicated that differential genes associated with cellular process and signaling were up-regulated in dispersed mycelia, besides significantly differential genes related to metabolism. The value of RPKM suggested that most of unique transcribed genes in both mycelia were transcribed at a low level, whereas those in the aggregated mycelia were more various and with higher value of RPKM than the dispersed one. Moreover, the higher lipase activity of aggregated R. chinensis agreed with the higher transcription level in the cells. [Conclusion] Our findings indicate intrinsic mechanisms of mycelia morphology differentiation and their influence on metabolism.