Abstract

Fungi play vital roles in ecosystems and biotechnology, yet synthetic biology efforts have predominantly focused on the model organism Saccharomyces cerevisiae, with less attention given to filamentous fungi. This review explores the molecular toolkits available for gene regulatory network (GRN) manipulation in these organisms, focusing on transcriptional rewiring and promoter engineering. Beyond constitutive and inducer-responsive promoters, the use of cis-elements responsive to physical cues like light and temperature enables gene expression control with unprecedented spatiotemporal precision. Additionally, newly available modular toolkits further facilitate GRN rewiring for metabolic engineering and fundamental research in filamentous fungi. Using Neurospora crassa as a model, we illustrate how transcriptional rewiring enhances cellulase production and reveals design principles of complex circuits, such as circadian oscillators. © 2025 Elsevier Ltd