Yumeng Chen1, Xingjia Fan1, Xinqing Zhao2, Yaling Shen1, Xiangyang Xu3, Liujing Wei1, Wei Wang4, Dongzhi Wei1. 1. State Key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, 130 Meilong Road , P.O.B. 311, Shanghai, 200237, China. 2. State Key Laboratory of Microbial Metabolism, Joint International Research Laboratory of Metabolic and Developmental Sciences, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. 3. Zaozhuang Jie Nuo Enzyme Co. Ltd., Shandong, China. 4. State Key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, 130 Meilong Road , P.O.B. 311, Shanghai, 200237, China. wadexp@ecust.edu.cn.
Abstract
BACKGROUND: The filamentous fungus Trichoderma reesei is one of the best producers of cellulase and has been widely studied for the production of cellulosic ethanol and bio-based products. We previously reported that Mn2+ and N,N-dimethylformamide (DMF) can stimulate cellulase overexpression via Ca2+ bursts and calcium signalling in T. reesei under cellulase-inducing conditions. To further understand the regulatory networks involved in cellulase overexpression in T. reesei, we characterised the Mn2+/DMF-induced calcium signalling pathway involved in the stimulation of cellulase overexpression. RESULTS: We found that Mn2+/DMF stimulation significantly increased the intracellular levels of cAMP in an adenylate cyclase (ACY1)-dependent manner. Deletion of acy1 confirmed that cAMP is crucial for the Mn2+/DMF-stimulated cellulase overexpression in T. reesei. We further revealed that cAMP elevation induces a cytosolic Ca2+ burst, thereby initiating the Ca2+ signal transduction pathway in T. reesei, and that cAMP signalling causes the Ca2+ signalling pathway to regulate cellulase production in T. reesei. Furthermore, using a phospholipase C encoding gene plc-e deletion strain, we showed that the plc-e gene is vital for cellulase overexpression in response to stimulation by both Mn2+ and DMF, and that cAMP induces a Ca2+ burst through PLC-E. CONCLUSIONS: The findings of this study reveal the presence of a signal transduction pathway in which Mn2+/DMF stimulation produces cAMP. Increase in the levels of cAMP activates the calcium signalling pathway via phospholipase C to regulate cellulase overexpression under cellulase-inducing conditions. These findings provide insights into the molecular mechanism of the cAMP-PLC-calcium signalling pathway underlying cellulase expression in T. reesei and highlight the potential applications of signal transduction in the regulation of gene expression in fungi.
BACKGROUND: The filamentous fungus Trichoderma reesei is one of the best producers of cellulase and has been widely studied for the production of cellulosic ethanol and bio-based products. We previously reported that Mn2+ and N,N-dimethylformamide (DMF) can stimulate cellulase overexpression via Ca2+ bursts and calcium signalling in T. reesei under cellulase-inducing conditions. To further understand the regulatory networks involved in cellulase overexpression in T. reesei, we characterised the Mn2+/DMF-induced calcium signalling pathway involved in the stimulation of cellulase overexpression. RESULTS: We found that Mn2+/DMF stimulation significantly increased the intracellular levels of cAMP in an adenylate cyclase (ACY1)-dependent manner. Deletion of acy1 confirmed that cAMP is crucial for the Mn2+/DMF-stimulated cellulase overexpression in T. reesei. We further revealed that cAMP elevation induces a cytosolic Ca2+ burst, thereby initiating the Ca2+ signal transduction pathway in T. reesei, and that cAMP signalling causes the Ca2+ signalling pathway to regulate cellulase production in T. reesei. Furthermore, using a phospholipase C encoding gene plc-e deletion strain, we showed that the plc-e gene is vital for cellulase overexpression in response to stimulation by both Mn2+ and DMF, and that cAMP induces a Ca2+ burst through PLC-E. CONCLUSIONS: The findings of this study reveal the presence of a signal transduction pathway in which Mn2+/DMF stimulation produces cAMP. Increase in the levels of cAMP activates the calcium signalling pathway via phospholipase C to regulate cellulase overexpression under cellulase-inducing conditions. These findings provide insights into the molecular mechanism of the cAMP-PLC-calcium signalling pathway underlying cellulase expression in T. reesei and highlight the potential applications of signal transduction in the regulation of gene expression in fungi.
Authors: Vijai Kumar Gupta; Andrei Stecca Steindorff; Renato Graciano de Paula; Rafael Silva-Rocha; Astrid R Mach-Aigner; Robert L Mach; Roberto N Silva Journal: Trends Biotechnol Date: 2016-07-06 Impact factor: 19.536