Synthetic Gene Network with Positive Feedback Loop Amplifies Cellulase Gene Expression in i>Neurospora crassa/i>

摘要

Second-generation or lignocellulosic biofuels are a tangible source of renewable energy, which is critical to combat climate change by reducing the carbon footprint. Filamentous fungi secrete cellulose-degrading enzymes called cellulases, which are used for production of lignocellulosic biofuels. However, inefficient production of cellulases is a major obstacle for industrial-scale production of second-generation biofuels. We used computational simulations to design and implement synthetic positive feedback loops to increase gene expression of a key transcription factor, CLR-2, that activates a large number of cellulases in a filamentous fungus, i>Neurospora crassa . Overexpression of CLR-2 reveals previously unappreciated roles of CLR-2 in lignocellulosic gene network, which enabled simultaneous induction of approximately 50% of 78 lignocellulosic degradation-related genes in our engineered i>Neurospora strains. This engineering results in dramatically increased cellulase activity due to cooperative orchestration of multiple enzymes involved in the cellulose degradation pathway. Our work provides a proof of principle in utilizing mathematical modeling and synthetic biology to improve the efficiency of cellulase synthesis for second-generation biofuel production.