BackgroundLignocellulose is the most abundant renewable resource in the world and has attracted widespread attention. It can be hydrolyzed into sugars with the help of cellulases and hemicellulases that are secreted by filamentous fungi. Several studies have revealed that the Ras small GTPase superfamily regulates important cellular physiological processes, including synthesis of metabolites, sporulation, and cell growth and differentiation. However, it remains unknown how and to what extent Ras small GTPases participate in cellulase production.ResultsIn this study, we found that the putative Ras small GTPase RSR1 negatively regulated the expression of cellulases and xylanases. Deletion ofrsr1(∆rsr1) significantly increased cellulase production and decreased the expression levels of ACY1-cAMP-protein kinase A (PKA) signaling pathway genes and the concentration of intracellular cyclic adenosine monophosphate (cAMP). Loss ofacy1based on ∆rsr1(∆rsr1∆acy1) could further increase cellulase production and the expression levels of cellulase genes, while overexpression ofacy1based on ∆rsr1(∆rsr1-OEacy1) significantly reduced cellulase production and transcriptional levels of cellulase genes. In addition, our results revealed that RSR1 negatively controlled cellulase production via the ACY1-cAMP-PKA pathway. Transcriptome analysis revealed significantly increased expression of three G-protein coupled receptors (GPCRs;tre62462,tre58767, andtre53238) and approximately two-fold higher expression of ACE3 and XYR1, which transcriptionally activated cellulases with the loss ofrsr1. ∆rsr1∆tre62462exhibited a decrease in cellulase activity compared to ∆rsr1, while that of ∆rsr1∆tre58767and ∆rsr1∆tre53238showed a remarkable improvement compared to ∆rsr1.These findings revealed that GPCRs on the membrane may sense extracellular signals and transmit them torsr1and then to ACY1-cAMP-PKA, thereby negatively controlling the expression of the cellulase activators ACE3 and XYR1. These data indicate the crucial role of Ras small GTPases in regulating cellulase gene expression.ConclusionsHere, we demonstrate that some GPCRs and Ras small GTPases play key roles in the regulation of cellulase genes inTrichoderma reesei. Understanding the roles of these components in the regulation of cellulase gene transcription and the signaling processes inT. reeseican lay the groundwork for understanding and transforming other filamentous fungi.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13068-023-02341-z.