Promoters and Synthetic Promoters in Trichoderma reesei.

ABSTRACT

Trichoderma reesei holds immense promise for large-scale protein production, rendering it an excellent subject for deeper exploration using genetic engineering methods to achieve a comprehensive grasp of its cellular physiology. Understanding the genetic factors governing its intrinsic regulatory network is crucial, as lacking this knowledge could impede the expression of target genes. Prior and ongoing studies have concentrated on advancing new expression systems grounded in synthetic biology principles. These methodologies involve utilizing established potent promoters or engineered variations. Genomic and transcriptomic analyses have played a pivotal role in identifying robust promoters and expression systems, including light-responsive, copper-inducible, L-methionine-inducible, and Tet-On systems, among others. This chapter seeks to highlight various research endeavors focusing on tunable and constitutive promoters, the impact of different promoters on both native and foreign protein expression, the discovery of fresh promoters, and strategies conducive to future research aimed at refining and enhancing protein expression in T. reesei. Characterizing new promoters and adopting innovative expression systems hold the potential to significantly expand the molecular toolkit accessible for genetically engineering T. reesei strains. For instance, modifying potent inducible promoters such as Pcbh1 by replacing transcriptional repressors (cre1, ace1) with activators (xyr1, ace2, ace3, hap2/3/5) and integrating synthetic expression systems can result in increased production of crucial enzymes such as endoglucanases (EGLs), ß-glucosidases (BGLs), and cellobiohydrolases (CBHs). Similarly, robust constitutive promoters such as Pcdna1 can be converted into synthetic hybrid promoters by incorporating activation elements from potent inducible promoters, facilitating cellulase induction and expression even under repressive conditions. Nevertheless, further efforts are necessary to uncover innovative promoters and devise novel expression strategies to enhance the production of desired proteins on an industrial scale.