Li Ni, Qiu Zhouyuan, Cai Wanchuan, Shen Yaling, Wei Dongzhi, Chen Yumeng, Wang Wei
The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, P.O.B. 311, Shanghai, 200237, China.
Jiangsu Yiming Biological Technology Co., Ltd., Suqian, 223699, Jiangsu, China.
Biotechnol Biofuels Bioprod. 2023 May 23;16(1):87. doi: 10.1186/s13068-023-02341-z.
Lignocellulose is the most abundant renewable resource in the world and has attracted widespread attention. It can be hydrolyzed into sugars with the help of cellulases and hemicellulases that are secreted by filamentous fungi. Several studies have revealed that the Ras small GTPase superfamily regulates important cellular physiological processes, including synthesis of metabolites, sporulation, and cell growth and differentiation. However, it remains unknown how and to what extent Ras small GTPases participate in cellulase production.
In this study, we found that the putative Ras small GTPase RSR1 negatively regulated the expression of cellulases and xylanases. Deletion of rsr1 (∆rsr1) significantly increased cellulase production and decreased the expression levels of ACY1-cAMP-protein kinase A (PKA) signaling pathway genes and the concentration of intracellular cyclic adenosine monophosphate (cAMP). Loss of acy1 based on ∆rsr1 (∆rsr1∆acy1) could further increase cellulase production and the expression levels of cellulase genes, while overexpression of acy1 based on ∆rsr1 (∆rsr1-OEacy1) significantly reduced cellulase production and transcriptional levels of cellulase genes. In addition, our results revealed that RSR1 negatively controlled cellulase production via the ACY1-cAMP-PKA pathway. Transcriptome analysis revealed significantly increased expression of three G-protein coupled receptors (GPCRs; tre62462, tre58767, and tre53238) and approximately two-fold higher expression of ACE3 and XYR1, which transcriptionally activated cellulases with the loss of rsr1. ∆rsr1∆ tre62462 exhibited a decrease in cellulase activity compared to ∆rsr1, while that of ∆rsr1∆tre58767 and ∆rsr1∆tre53238 showed a remarkable improvement compared to ∆rsr1. These findings revealed that GPCRs on the membrane may sense extracellular signals and transmit them to rsr1 and then to ACY1-cAMP-PKA, thereby negatively controlling the expression of the cellulase activators ACE3 and XYR1. These data indicate the crucial role of Ras small GTPases in regulating cellulase gene expression.
Here, we demonstrate that some GPCRs and Ras small GTPases play key roles in the regulation of cellulase genes in Trichoderma reesei. Understanding the roles of these components in the regulation of cellulase gene transcription and the signaling processes in T. reesei can lay the groundwork for understanding and transforming other filamentous fungi.
木质纤维素是世界上最丰富的可再生资源，已引起广泛关注。借助丝状真菌分泌的纤维素酶和半纤维素酶，它可以水解成糖。多项研究表明，Ras小GTP酶超家族调节重要的细胞生理过程，包括代谢物合成、孢子形成以及细胞生长和分化。然而，Ras小GTP酶如何以及在多大程度上参与纤维素酶的产生仍不清楚。
在本研究中，我们发现推定的Ras小GTP酶RSR1负向调节纤维素酶和木聚糖酶的表达。rsr1缺失（∆rsr1）显著增加了纤维素酶的产量，并降低了ACY1-环磷酸腺苷-蛋白激酶A（PKA）信号通路基因的表达水平和细胞内环磷酸腺苷（cAMP）的浓度。基于∆rsr1的acy1缺失（∆rsr1∆acy1）可进一步提高纤维素酶产量和纤维素酶基因的表达水平，而基于∆rsr1的acy1过表达（∆rsr1-OEacy1）则显著降低纤维素酶产量和纤维素酶基因的转录水平。此外，我们的结果表明RSR1通过ACY1-cAMP-PKA途径负向控制纤维素酶的产生。转录组分析显示，三种G蛋白偶联受体（GPCRs；tre62462、tre58767和tre53238）的表达显著增加，且ACE3和XYR1的表达增加了约两倍，随着rsr1缺失，它们转录激活纤维素酶。与∆rsr1相比，∆rsr1∆tre62462的纤维素酶活性降低，而∆rsr1∆tre58767和∆rsr1∆tre53238的纤维素酶活性与∆rsr1相比有显著提高。这些发现表明，膜上的GPCRs可能感知细胞外信号并将其传递给rsr1，然后传递给ACY1-cAMP-PKA，从而负向控制纤维素酶激活剂ACE3和XYR1的表达。这些数据表明Ras小GTP酶在调节纤维素酶基因表达中起关键作用。
在此，我们证明了一些GPCRs和Ras小GTP酶在里氏木霉纤维素酶基因的调控中起关键作用。了解这些成分在里氏木霉纤维素酶基因转录调控和信号传导过程中的作用，可为理解和改造其他丝状真菌奠定基础。
