A light-switchable bidirectional expression system in filamentous fungus Trichoderma reesei.
The filamentous fungi Trichoderma reesei is widely used in the production of cellulolytic enzymes and recombinant proteins. However, only moderate success has been achieved in expressing heterologous proteins in T. reesei. Light-dependent control of DNA transcription, and protein expression have been demonstrated in bacteria, fungi, and mammalian cells. In this study, light inducible transactivators, a "light-on" bidirectional promoter and a "light-off" promoter were constructed successfully in T. reesei for the first time. Our light inducible transactivators can homodimerize and bind to the upstream region of artificial promoters to activate or repress genes transcription. Additionally, we upgraded the light-inducible system to on-off system that can simultaneously control the expression of multiple heterologous proteins in T. reesei. Moreover, a native cellulase-free background for the expression of heterologous proteins was achieved by knocking out the genes involved in transcriptional regulation and encoding of cellulases: xyr1, cbh1, and cbh2. Our light-switchable system showed a very little background protein expression and robust activation in the blue light with significantly improved heterologous protein expression. We demonstrate that our light-switchable system has a potential application as an on/off "switch" that can simultaneously regulate the expression of multiple genes in T. reesei under native cellulase-free background.
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Guoxiu
G
Zhang
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Pei
P
Liu
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Wei
W
Wei
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Xuedong
X
Wang
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China.
Dongzhi
D
Wei
State key Lab of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, China. Electronic address: wadexp@ecust.edu.cn.
Wei
W
Wang
D015966:Gene Expression Regulation, Fungal; D049452:Green Fluorescent Proteins; D008027:Light; D008828:Microbiological Techniques; D008856:Microscopy, Fluorescence; D011401:Promoter Regions, Genetic; D060888:Real-Time Polymerase Chain Reaction; D014242:Trichoderma / Q000235:genetics*
D049452:Green Fluorescent Proteins
Cellulase-free background; Heterologous protein; Light-switchable system; Simultaneous expression; Trichoderma reesei